Extracts of Isochrysis sp.

ABSTRACT

The present invention relates to extracts of Isochrysis sp., preferably Tahitian Isochrysis, its cosmetic, dermatological and/or therapeutic uses and compositions and cosmetic, dermatological or therapeutic products comprising such an extract of Isochrysis sp., preferably Tahitian Isochrysis.

CROSS-REFERENCE TO RELATED APPLICATION

This application claims benefit of priority to U.S. ProvisionalApplication No. 61/101,228, filed on Sep. 30, 2008, which isincorporated herein by reference in its entirety.

The present invention relates to extracts of Isochrysis sp., preferablyTahitian Isochrysis, its cosmetic, dermatological and/or therapeuticuses and compositions and cosmetic, dermatological or therapeuticproducts comprising such an extract of Isochrysis sp., preferablyTahitian Isochrysis.

In the field of cosmetic and dermatology, there exists a need to provideagents for influencing or modifying the growth of human hair, i.e. toprovide agents for increasing the speed of human hair growth and/or forstimulating hair growth from otherwise non-productive hair follicles.Likewise, agents for influencing or modifying growth of human hair thatreduce the speed of growth of human hair and/or for unhairing aresought. Preferably, the influencing or modifying of the growth of humanhair should be locally confinable.

On the other hand, in the field of cosmetics there are sought agents forinfluencing or modifying pigmentation of human skin and/or hair, i.e.agents for increasing coloration of human skin and/or hair (hereinafteralso called “browning” or “tanning”) or for decreasing pigmentation ofthe skin and/or hair (hereinafter also called “lightening”).

Hair growth is not a continuous, steadily ongoing procedure, but insteadresults from the production of hair material in hair follicles thatundergo several stages of a hair growth cycle. During a rest phase ofthe hair follicle, no significant amount of hair material is produced,while during a hair Eco production phase the production of hair materialis started or continued. Some hair follicles turn into an apparentlydormant state, in which for an apparently indefinite time no furtherhair material is produced, resulting in a loss of hairs.

Genetic disposition as well as the natural aging process and/or diseasecontribute to hair loss and slower hair growth in both males andfemales. Approximately 50% of the population displays this trait to somedegree by the age of 50, where thinning of the hair can begin between 12and 40 years of age independent of gender (Otberg N et al., EndocrinolMetab Clin North Am. 2007, 36(2), 379-398 and Price V H., InvestigDermatol Symp Proc. 2003, 8 (1), 24-27).

To increase the growth of human hair it has thus been proposed toprolong the phase of production of hair material and/or to shorten theresting phase of hair follicles, e.g. by reactivating apparently dormanthair follicles. Thus, agents able to stimulate hair growth as well asprevent and slow down or reduce hair loss could be beneficial not onlyas a cure for alopecia but also as positively affect the psychosocialevents associated with hair disorders. Studies reveal psychosocialimpact with hair loss to include body image dissatisfaction associatedwith negative stereotypes such as feeling older, weaker and lessattractive (S. Pickard-Holley, Sem. Oncol. Nurs. 1995, 11, 235-238).

Various agents for stimulating hair growth have been proposed. Drugs,including Minoxidil (Rogaine), Finasteride (Propecia) and Dutasteride(Avodart) are approved treatments for hair loss. However, they requiremedical prescription, and are active only on a certain percentage of thepopulation. Moreover, some of these drugs are not permitted to be usedby females because of hormonal effects. Thus, premenopausal women shouldnot take Finesteride due to the risk of abnormalities in male fetus whenbecoming pregnant (S. Krus et al., J. Appl. Cosmetol. 2007, 25, 59-74).

Minoxidil is a drug that is effective in inducing hair growth for asmall percentage of patients and will re-grow hair only on top of thescalp. Adverse effects when taken orally are tachycardia, anginapectoris and fluid retention. When applied topically adverse effects aremainly dermatologic, i.e. local irritation, itching, dryness anderythema (S. Krus et al., J. Appl. Cosmetol. 2007, 25, 59-74).

Other medical treatments available to treat hair loss include drasticsurgical techniques such as scalp reduction, scalp flaps or follicularunit transplantation. These surgeries carry the risk of complicationssuch as elevation of hairline associated with donor region, possibilityof necrosis and unnatural appearance of hair growth direction,anesthesia and post-op care, not to mention high costs.

Herbal preparations that claim to induce hair growth (e.g. Hair Prime)are available at low cost but their effectiveness is often very limited.

On the other hand, it is sometimes desirable to unhair parts of a humanbody. Thus, it is for example generally preferred to have hair on thescalp but very often hair is unwanted on other parts of the body,especially on the legs, under the arms and on the face. Furthermorethere are pathological hair growth disorders (e. g. hirsutism,folliculitis, pseudofolliculitis barbea) that require medical treatment.

Various procedures have been employed to remove unwanted hair, includingshaving, electrolysis, depilatory creams or lotions, waxing, plucking,and therapeutic antiandrogens. These conventional procedures generallyhave drawbacks associated with them. Shaving, for instance, can causenicks and cuts, and can leave a perception of an increase in the rate ofhair regrowth. Shaving also can leave an undesirable stubble.Electrolysis, on the other hand, can keep a treated area free of hairfor prolonged periods of time, but can be expensive, painful, andsometimes leaves scarring. Depilatory creams, though very effective,typically are not recommended for frequent use due to their highirritancy potential. Waxing and plucking can cause pain, discomfort, andpoor removal of short hair. Finally, antiandrogens, which have been usedto treat female hirsutism, can have unwanted side effects.

Skin- or hair-lightening active ingredients intervene in one form oranother in melanin metabolism or catabolism. Melanin pigments, which arenormalty brown to black in colour, are formed in the melanocytes of theskin, transferred to the keratinocytes and give the skin or hair itscolour. In mammals, the brown-black eumelanins are primarily formed fromhydroxysubstituted aromatic amino acids such as L-tyrosine and L-DOPA,the yellow to red pheomelanins additionally from sulfur-containingmolecules (Cosmetics & Toiletries 1996, 111 (5), 43-51). Starting fromL-tyrosine, L3,4-dihydroxyphenylalanine (L-DOPA) is formed by thecopper-containing key enzyme tyrosinase and is in turn converted bytyrosinase to dopachrome. By a series of steps catalysed by variousenzymes, the latter is oxidised to form melanin.

Skin-lightening agents are used for various reasons: if for some reasonthe melanin-forming melanocytes in human skin are not evenlydistributed, pigment spots occur which are either lighter or darker thanthe surrounding skin area. To overcome this problem, skin and hairlightening agents are sold which at least partially help to balance outsuch pigment spots. In addition, many people have a need to lightentheir naturally dark skin colour or to prevent skin pigmentation.Hair-lightening agents are useful to lighten regrowing unwanted hair andmake it thus less noticeable. In addition, many people have the desireto lighten their naturally dark hair colour. This requires very safe andeffective skin and hair lightening agents. Many skin and hair lighteningagents contain more or less powerful tyrosinase inhibitors. This is onlyone possible route towards skin and hair lightening, however.

Furthermore, UV-absorbing substances are also used to protect againstthe increase in skin pigmentation caused by UV light. This is a purelyphysically induced effect, however, and must be distinguished from thebiological action of skin-lightening agents on cellular melaninformation, which can also be detected in the absence of UV light.Moreover, UV absorbers do not bring about a true lightening of the skinbut merely inhibit the increase in skin pigmentation caused by UV light.

Hydroquinone, hydroquinone derivatives such as e.g. arbutin, vitamin C,derivatives of ascorbic acid such as e.g. ascorbyl palmitate, kojic acidand derivatives of kojic acid such as e.g. kojic acid dipalmitate, areused in particular in commercial cosmetic or therapeutic skin and hairlightening 2.5 formulations.

One of the most commonly used skin and hair lighteners is hydroquinone.However, this compound has a cytotoxic effect on melanocytes and isirritating to the skin. For that reason such preparations are no longerauthorised for cosmetic applications in Europe, Japan and South Africa,for example. In addition, hydroquinone is very sensitive to oxidationand can be stabilised only with difficulty in cosmetic formulations.Arbutin is a hydroquinone glucoside, which hydrolyses in situ to formhydroquinone and is therefore just as questionable in toxicologicalterms as hydroquinone.

Vitamin C and ascorbic acid derivatives have only an inadequate effecton the skin. Furthermore, they do not act directly as tyrosinaseinhibitors but instead reduce the coloured intermediate stages ofmelanin biosynthesis.

Kojic acid (5-hydroxy-2-hydroxymethyl-4-pyranone) is a tyrosinaseinhibitor which inhibits its catalytic action by chelating the copperatoms in the enzyme; it is used in commercial skin and hair lighteningagents but has a high sensitising potential and causes contactallergies.

On the other hand, it is sometimes desirable to tan parts of the body.Skin and hair browning agents can at least partially help to balance outpigment spots if the melanin-forming melanocytes are not evenlydistributed. In addition, many people need to tint their naturally paleskin colour and to develop skin pigmentation without being exposed tosolar radiation. In addition, some people have the desire to obtain amore intense and homogeneous hair colour. For this reason very safe andeffective skin and hair browning agents are necessary.

It is also known that in fair-skinned people high exposure to the suncan cause the breakdown of the vitally important B vitamin folic acid.Folic acid deficiency in pregnancy for example leads to severedeformities. Folic acid is also necessary for DNA synthesis and istherefore essential for sperm production. Folic acid deficiency cantherefore lead to infertility. A protection against UV radiationaccordingly prevents folic acid deficiency.

Artificial skin browning can be carried out cosmetically or medically,the following main approaches playing a part:

If carotene preparations are taken regularly, carotene is stored in thefatty tissue of the subcutis and the skin gradually turns orange toyellow-brown.

Washable makeup preparations can be used to achieve a light skin tinting(e.g. extracts of fresh green walnut shells, henna).

Skin browning can also be achieved by chemical changes to the skin'sstratum corneum using so-called self-tanning preparations. The mostimportant active ingredient is dihydroxyacetone. The skin browningachieved in this way does not wash off and is only removed with thenormal flaking of the skin (after around 5 to 10 days). Dihydroxyacetonecan be classed as a ketotriose and as a reducing sugar it reacts withthe amino acids in the skin or the free amino and imino groups inkeratin via a series of intermediate steps along the lines of a Maillardreaction to form browncoloured substances known as melanoids, which areoccasionally also called melanoidins.

One disadvantage of this is that unlike “sun-tanned” skin, the skinbrowning obtained with dihydroxyacetone does not protect the skinagainst sunburn. A further disadvantage of dihydroxyacetone lies in thefact that, particularly under the influence of ultraviolet radiation, itreleases formaldehyde, albeit usually in small amounts. Dihydroxyacetonealso has an unpleasant, chemical odour.

The tint obtained with self-tanning agents is achieved without exposureto sunlight. In contrast, so-called “pre-tan products” or “tanpromoters” are also available, which have to be applied before exposureto sunlight. In the sun these preparations then turn yellow, giving riseto a light brown-yellow colouring of the epidermis which further booststhe “suntan”.

Another type of artificial browning which is not dependent on UV lightcan be brought about through the hormones which are usually alsoreleased in the body as a consequence of (natural) UV irradiation andultimately stimulate the melanocytes to synthesise melanin. Exampleswhich can be cited in this connection are derivatives ofproopiomelanocortin (POMC) such as [alpha]-MSH (Melanocyte StimulatingHormone) and synthetic variants (such as [Nle(4),D-Phe(7)]-[alpha]-MSH), which in some cases display far higher activitylevels than the natural [alpha]-MSH. Although these hormones can causebrowning in principle, their use in cosmetics is prohibited, since theyare pharmacologically potent substances (hormones) which should not bewidely used without medical indications. There exists thus in the art anongoing need to provide further agents for influencing or modifyinggrowth of human hair and/or pigmentation of human skin and/or hair.

According to the invention, there is thus provided a method of obtaininga composition for influencing or modifying

a) growth of human hair and/orb) pigmentation of human skin and/or hair,comprising the step of extracting cell material of Isochrysis sp.,preferably Tahitian Isochrysis, with a liquid extractant selected fromthe group consisting of hexane, ethyl acetate, ethanol, water, methanol,isopropanol and mixtures of two or more of these extractants,wherein the extraction comprises a) exposition of the cell material tothe extractant for up to 24 h at a temperature of not more than 50° C.,and b) removal of the cell material to obtain an extract, the extractbeing the composition or being further processed into the composition.

Microalgae have been used in the field of cosmetics before. For example,EP 1 745 794 A2 discloses the use of microalgae or microalgae extractsfor preventing or treating protozoal infections in human or animals.Also, U.S. Pat. No. 5,767,095 discloses topical anti-inflammatorycompositions comprising monogalactosyl-dieicosapentanoyl glycerolobtained from Isochrysis galbana. Whole cells of Isochrysis galbana havebeen used in FR 2 801 788 and FR 2 676 454 for achieving a stabilizedredox potential and for reconstituting of abiotic sea water forthalassotherapy. Also, several documents like EP 1 886 679, EP 918 517B1 and WO 94/24984 describe processes for obtaining specific substancesby extraction of microalgae cells. None of the documents teaches orsuggests the usefulness of Isochrysis and particularly of TahitianIsochrysis for influencing or modifying the growth of human hair and/orpigmentation of human skin and/or hair.

US 2006/269497 discloses the use of astaxanthin as a trichogenous agentsand also indicates that astaxanthin can be found in microalgae.Isochrysis and particularly Tahitian Isochrysis is not mentioned.

JP 2002-068943 A generally discloses uses of extracts or severalmicroalgae species and genera for inhibitingtestosterone-5-alpha-reductase. The order Isochrysis is generallymentioned. However, the document also mentions that special culturing isnecessary for obtaining microalgae cells, and different culturalconditions are required for each microalgae species. The document doesnot disclose culturing conditions for genus Isochrysis.

FR 2657012 A1 discloses in example 8 an extract of Isochrysis algaeobtained by introducing the algae in aqueous ethanol (water:ethanol=19:1(v/v)) and disintegrating said algae with a mixer having high shearforces (Ultra Turrax). After disintegration, extraction was carried outfor 24 hours at 20° C. The disrupted algae were subsequently separatedoff by filtration. In contrast to this, in the context of the presentinvention the microalgae are not disintegrated before extraction.Instead, the cell material is substantially or completely intact orfreeze-dried. Extracts obtained or obtainable from disrupted Isochrysisalgae clearly differ from those obtained in accordance with the presentinvention. According to the present invention cell material isconsidered substantially or completely intact if the cell membrane of90% of cells of a sample comprising 100 or more cells is intact,preferably as determined by propidium iodide staining and opticalinspection.

Thus, in the context of the present invention the “cell material ofIsochrysis sp.” and particularly “cell material of Tahitian Isochrysis”refers to a composition of freeze-dried, substantially or completelyintact cells or mixtures thereof, wherein the cells are Isochrysis sp.cells or Tahitian Isochrysis cells, respectively. The cell material cancomprise a carrier medium, provided that the total content of disruptedcells is less than 10% of all cells, preferably as determined aspropidium iodide staining. In summary, what is extracted according tothe present invention is not a homogenized or substantially disruptedmass of cells. Instead, the cell material according to the presentinvention is preferably obtained by a method consisting of the followingsteps:

-   -   1. Cultivating Isochrysis sp. cells and/or preferably of        Tahitian Isochrysis    -   2. Harvesting the cells to obtain completely or substantially        intact cell material,    -   3. Optionally washing the cell material of step 2 once or        multiple times, to obtain washed, substantially or completely        intact cell material,    -   4. Optionally freeze-drying the cell material of step 2 and/or        step 3.

It is generally known that different biological species comprisedifferent substances. Thus, effects obtainable by use of one microalgalspecies cannot be used to predict the effects obtainable by use of adifferent microalgal species. Furthermore, as will be shown hereinafter,the effects obtainable by extracts of Isochrysis sp., preferablyTahitian Isochrysis, largely depend on the exact extraction conditionsand may even be reversed upon modification of the extraction methodused.

It was thus surprising that extracts of Isochrysis sp., preferablyTahitian Isochrysis, are useful for influencing or modifying growth ofhuman hair and/or pigmentation of human skin and/or hair. Also, theextracts of the present invention favourably allow the production ofeffective compositions by simple and reliable methods starting frombiological materials, i.e. microalgal cell material.

Isochrysis sp., preferably Tahitian Isochrysis, is employed forobtaining extracts and compositions according to the present invention.Tahitian Isochrysis is a strain of Isochrysis collectable at Mataiva(Tahiti), According to the present invention, strain CS 177 obtainablefrom the Australian CSIRO collection (also registered as CCMP1324 atProvasoli-Guillard National Center for Culture of Marine; NEPCC601 atthe Canadian Center for the Culture of Microorganisms) is preferablyused. This strain has been isolated by K. Haines in 1977 at Mataiva,Tahiti.

According to the present invention, cell material of Isochrysis sp.,preferably Tahitian Isochrysis, is extracted with a liquid extractantselected from the group consisting of hexane, ethyl acetate, ethanol,water, methanol. The extractant can also be a mixture of two or more ofthe aforementioned extractants such as for example a mixture ofhexane/ethyl acetate 1:1 (v/v). These extractants have provided bestresults for influencing or modifying growth of human hair and/orpigmentation of human skin and/or hair.

For extraction, the cell material is in step a) contacted with theliquid extractant for up to 24 h at a temperature of not more than 50°C., preferably at a temperature of 16-40° C. and most preferably at atemperature of 20-30° C. Also, exposition of the cell material to theextractant preferably lasts for up to 24 h, more preferably for 1-10 hand most preferably for 2-6 h. For all extraction conditions describedherein, best results have been achieved when the extraction wasperformed in the dark. Also, during contact of the cell material withthe extractant the material is preferably agitated, preferably bystirring.

Preferably, the ratio (w:v) of cell material to liquid extractant is 200mg: 1 ml to 1 mg:1 ml, more preferably 140 mg:1 ml to 5 mg:1 ml, andmost preferably 80 mg:1 ml to 10 mg:1 ml when using freeze-dried cellmaterial as detailed below.

After extraction, in step b) an extract is obtained by removal of thecell material, preferably by centrifugation, filtration or decantationor other suitable methods. A particle-free supernatant according tovisual inspection is thus obtained, typically of yellow-greenish tobrown-greenish colour. The extract can be used as a composition forinfluencing or modifying growth of human hair and/or pigmentation ofhuman skin and/or hair, or, more preferably, is further processed intosuch composition as detailed below.

The cell material removed from the extract in step b) can be used foranother exposition to the extractant in step a), typically for a fewminutes, preferably up to 1 h. The extraction thus preferably comprisesrepeating steps a) and b) once, twice, three or four times, preferablyonce or twice, and in each step a) the cell material removed in therespective prior step b) is used, and the extracts of steps b) arecombined. This way, a continued extract with reproducible compositionand high yield of extracted active ingredients can be obtained.

In a preferred method according to the present invention, the cellmaterial used in a step a) of the extraction is obtained in step b) of aprior extraction with a different extractant. Thus, two extracts orcompositions are provided, and the method can be repeated to providefurther extracts or compositions.

Particularly preferred are extracts and compositions obtainable orobtained by the following extractions:

1. Extraction with ethyl acetate, followed by extraction with ethanol,followed by extraction with water or extraction with ethyl acetatefollowed by extraction with 30% aqueous ethanol or with water2. Extraction with methanol or ethanol, followed by extraction withwater3. Extraction with hexane, followed by extraction with ethyl acetate,followed by extraction with ethanol, followed by extraction with water.4. Extraction with methanol or ethanol, followed by extraction with amixture of hexane/ethyl acetate 1:1 (v/v), hexane or ethyl acetatefollowed by extraction with water

The phototoxicity of the extracts and composition is preferably adjustedto a phototoxicity index of less than 5 according to the OfficialJournal of the European Communities, directive 2000/33/EG of thecommission from Apr. 25, 2000, appendix II, B.41 and the OECD Guideline432. At a phototoxicity index of less than 5, the composition is nolonger considered to possess phototoxic potential.

For adjusting phototoxicity the optionally combined extract—not in solidform—is preferably treated with activated carbon in a ratio dryextract:activated carbon of 3:1 to 1:20 (w/w), preferably 1:1 to 1:12(w/w). The term ‘dry extract’ according to the present invention refersto the extractant-free weight of an extract according to the presentinvention, e.g. as obtainable by completely removing the extractant.This treatment may result in a minor loss of activity for influencing ormodifying the growth of human hair and/or pigmentation of human skinand/or hair. However, the treatment reliably removes phototoxicingredients otherwise comprised in the extract. A further benefit ofthis treatment is a colour reduction of most of the extracts.

For preparing the composition, the extractant is preferably removed fromthe—optionally combined—extract, preferably by evaporation or other 1C)suitable processes, to obtain the extract in concentrated or dry form(dry extract), the latter preferably as a suspension, a viscous liquid,a powder or as granules. The concentrated form preferably contains 50-80wt. % dry matter and 50-20 wt. % residual extractant.

The composition is then formed by optionally adding a cosmeticallyand/or dermatologically and/or therapeutically acceptable solid carrierto the extract in concentrated or dry form (dry extract) and thenoptionally drying the mixture by suitable processes. In this context,such a solid which is at least not toxic to the organisms on which it isto be used is cosmetically, dermatologically or therapeuticallyacceptable. Preferred solids are hydrocolloids such as starches,degraded or chemically or physically modified starches (in particulardextrins and maltodextrin), lactose, modified celluloses, gum arabic,gum ghatti, tragacanth gum, karaya, carrageenan, pullulan, curdlan,xanthan gum, gellan gum, guar gum, locust bean gum, alginates, agar,pectin, inulin or glucose and mixtures of two or more of these solids.

The composition can also be formed by optionally adding a cosmeticallyand/or dermatologically and/or therapeutically acceptable solvent, suchas e.g. neutral oil, mineral oil, silicone oil, plant oils,triglycerides, fatty alcohols, fatty acid esters, polyol fatty acidesters such as PEG-7 glyceryl cocoate available e.g. as Cetiol HE fromCognis, ethanol, 1,2-propylene glycol, 1,3-butylene glycol,dipropyleneglycol, triethyl citrate, 1,2-pentanediol or other1,2-alkanediols, glycerin and water and mixtures of two or more of thesesolvents to the extract in concentrated or dry form (dry extract) andoptionally completely removing the residual extractant by a suitableprocess. Such compositions prepared according to the invention arereadily further processable in particular for cosmetic purposes. Thesecompositions can optionally be prepared with the addition of asolubilizing agent, preservative or antioxidant.

Most preferably, 1,2-pentanediol or 1,2-hexanediol or a mixture of oneof these dials and one or more of the above mentioned solvents forexample a mixture of water and 1,2-diol is selected as solvent. Thedials not only possess very good solubilizing properties but exhibitalso bioavailability enhancing and moisturizing activity. Furthermore,depending on the concentration of the 1,2-diol no further preservationor only reduced levels of preservatives are needed to protect thecomposition from microbial growth. Antioxidants such as for exampletocopherol or tocopherol mixtures, tocopherol acetate, BHT or othersuitable antioxidants are also easily incorporated.

For very lipophilic extracts such as for example hexane or ethyl acetateextracts, neutral oil, mineral oil, silicone oil, plant oils,triglycerides, fatty alcohols, fatty acid esters, polyol fatty acidesters, dipropylenglycol, triethyl citrate and ethanol and mixtures oftwo or more of these solvents are preferred.

The extract or the liquid or solid composition comprising the extractcan furthermore also be further processed by encapsulation with a solidshell material, which is preferably chosen from starches, degraded orchemically or physically modified starches (in particular dextrins andmaltodextrins), gelatines, wax materials, liposomes, gum arabic,agar-agar, ghatti gum, gellan gum, modified and non-modified celluloses,pullulan, curdlan, carrageenans, algic acid, alginates, pectin, inulin,xanthan gum and mixtures of two or more of the substances mentioned.

The solid shell material is preferably selected from gelatine (pork,beef, poultry and/or fish gelatines and mixtures thereof areadvantageous, preferably including at least one gelatine having a Bloomvalue of greater than or equal to 200, preferably having a Bloom valueof greater than or equal to 240), maltodextrin (preferably obtained frommaize, wheat, tapioca or potato, preferred maltodextrins displaying a DEvalue in the range from 10 to 20), modified cellulose (e.g. celluloseether), alginates (e.g. Na alginate), carrageenan (beta-, iota-, lambda-and/or kappa-carrageenan), gum arabic, curdlan and/or agar-agar.Gelatine is used in particular because of its good availability invarious Bloom values. Production can take place as described for examplein EP 0 389 700 A, JP 7 196 478, U.S. Pat. Nos. 4,251,195, 6,214,376, WO03/055587 or WO 2004/050069.

The compositions in liquid, solid (other than the dry extract) orencapsulated form obtainable or obtained according to the presentinvention comprise 0.001 to 20 wt. %, preferably 0.01 to 10 wt % andmost preferably 0.1-5 wt % dry extract relative to the totalcomposition.

According to the invention, a composition for stimulating growth ofhuman hair without or with only low activity in influencing pigmentationof human skin and/or hair is provided, the composition being orcomprising an extract obtained by using methanol as extractant. It hassurprisingly been found that by directly extracting Isochrysis sp.,preferably Tahitian Isochrysis, cell material, particularly freeze-driedmaterial, with methanol, the aforementioned effects can be achieved.

Further according to the invention, a composition for stimulating growthof human hair and increasing pigmentation of human skin and/or hair isprovided, wherein the composition is or comprises an extract obtained byusing ethyl acetate as extractant. It has surprisingly been found thatby directly extracting Isochrysis sp., preferably Tahitian Isochrysis,cell material, particularly freeze-dried material, with ethyl acetate,the aforementioned effects can be achieved.

Furthermore according to the invention, a composition for stimulatinggrowth of human hair and increasing pigmentation of human skin and/orhair is provided, wherein the composition is or comprises an extractobtained by using a hexane/ethyl acetate 1:1 (v/v) mixture after firstextracting the biomass with methanol as extractant. It has surprisinglybeen found that by extracting Isochrysis sp., preferably TahitianIsochrysis, cell material, particularly freeze-dried material, withmethanol followed by extraction with hexane/ethyl acetate 1:1 (v/v), theaforementioned effects can be achieved. A further benefit of thisextract is the very light yellowish colour. Also according to theinvention, a composition for increasing pigmentation of human skinand/or hair without or with only low activity in stimulating growth ofhuman hair, wherein the composition is or comprises an extract obtainedby using water as extractant. It has surprisingly been found that byextracting Isochrysis sp., preferably Tahitian Isochrysis, cell materialwith water, preferably after prior extraction of freeze-dried materialwith methanol, ethanol, ethyl acetate and/or hexane or mixtures of twoor more of these solvents, the aforementioned effects can be achieved.

For inhibiting growth of human hair while increasing pigmentation ofhuman skin and/or hair, a composition is provided according to theinvention that is or comprises an extract obtained by using ethanol asextractant on cell material obtained after extraction with ethylacetateor with hexane followed by extraction with ethylacetate. Againsurprisingly, such composition allows to achieve the aforementionedeffects.

Also according to the invention there is provided a composition forinhibiting growth of human hair and decreasing pigmentation of humanskin and/or hair, wherein the composition is or comprises an extractobtained by using ethanol as extractant. It has surprisingly been foundthat by directly extracting Isochrysis sp., preferably TahitianIsochrysis, cell material, particularly freeze-dried material, withethanol, the aforementioned effects can be achieved.

The composition of the present invention can favourably be part of acosmetic, dermatologic or therapeutic product. In such product thecomposition is preferably present in an amount sufficient to achieve theaforementioned effects upon application of the product to the human skinand/or hair or after oral consumption.

The concentration of the composition in a cosmetic, dermatological ortherapeutic product (for topical or oral application) preferably is

-   -   at least 0.001 ppm, preferably at least and 0.01 ppm and most        preferably at least 0.1 ppm, and    -   at most 100 ppm, preferably at most 50 ppm and most preferably        at most 10 ppm dry extract of the total product.

The cosmetic, dermatological or therapeutic products according to theinvention are produced by conventional processes known per se, such thatthe extract or the extract composition is incorporated into cosmetic,dermatological or therapeutic products which can have a conventionalcomposition and which in addition to the aforementioned effects can alsobe used for the treatment, care and cleansing of the skin or hair.

Essential fields of use for extract or extract compositions according tothe invention are cosmetic, dermatological or therapeutic products which(apart from the presence of the extract according to the invention)serve for cosmetic or dermatological light protection, for treatment,care and cleansing of the skin and/or hair or as a make-up product indecorative cosmetics. Such products can accordingly be present e.g. as acleansing composition, such as e.g. soap, syndet, liquid washing, showerand bath preparation, skin care composition, such as e.g. emulsion (as asolution, dispersion, suspension; cream, lotion or milk of the W/O, O/Wor multiple emulsion, PIT emulsion, emulsion foam, micro- ornanoemulsion, Pickering emulsion type, depending on the preparationprocess and constituents), ointment, paste, gel (including hydro-,hydrodispersion-, oleogel), alcoholic or aqueous/alcoholic solution,oil, toner, balsam, serum, powder (e.g. face powder, body powder),soaking liquid for wipes, Eau de Toilette, Eau de Cologne, perfume, wax,including the presentation form as a mask, mousse, stick, pencil,roll-on, (pump) spray, aerosol (foaming, non-foaming or after-foaming),skin care composition (as described above) as a foot care composition(including keratolytics, deodorant), as an insect repellent composition,as a sunscreen composition, as a self-tanning composition and/oraftersun preparation, skin care composition as a shaving composition orafter-shave, as a hair-removing composition, as a hair care composition,such as e.g. shampoo (including shampoo for normal hair, for greasyhair, for dry, stressed (damaged) hair, 2-in-1 shampoo, anti-dandruffshampoo, baby shampoo, shampoo for a dry scalp, shampoo concentrate),conditioner, hair treatment cure, hair tonic, hair lotion, hair rinse,styling cream, pomade, permanent wave and fixing compositions, hairsmoothing composition (straightening composition, relaxer), hair settingcomposition, styling aid (e.g. gel or wax); blonding composition, haircolouring composition, such as e.g. temporary, directly absorbed,semi-permanent hair colouring composition, permanent hair colouringcomposition), skin care composition as a decorative body carecomposition, such as e.g. nail care composition (nail varnish and nailvarnish remover), decorative cosmetic (e.g. powder, eye shadow, kajalpencil, lipstick, mascara), make-up, make-up remover, skin carecomposition as a deodorant and/or antiperspirant.

It is also advantageous to administer the extract or extract compositionorally e.g. in the form of tablets, dragees, capsules, juices, solutionsand granules or in form of orally consumable products used foralimentation which in addition to their function as foodstuff providebeauty from inside.

Compositions according to the present invention can advantageously becombined, in particular in cosmetic products, with further conventionalcomponents, such as, for example:

preservatives, in particular those described in US 2006/0089413,antimicrobial agents, such as e.g. antibacterial agents or agents totreat yeast and mold, in particular those described in WO 2005/123101,antiacne and sebum reducing agents, in particular those described in WO2008/046791, compounds against ageing of the skin, in particular thosedescribed in WO 2005/123101, antidandruff agents, in particular thosedescribed in WO 2008/046795, antiirritants (antiinflammatory agents,irritation-preventing agents, irritation-inhibiting agents), inparticular those described in WO 2007/042472 and US 2006/0089413,antioxidants, in particular those described in WO 2005/123101, carriermaterials, in particular those described in WO 2005/123101, chelatingagents, in particular those described in WO 2005/123101, deodorizingagents and antiperspirants, in particular those described in WO2005/123101, moisture regulators (moisture-donating agents, moisturizingsubstance, moisture-retaining substances), in particular those describedin WO 2005/123101, osmolytes, in particular those described in WO2005/123101, compatible solutes, in particular those described in WO01/76572 and WO 02/15868, proteins and protein hydrolysates, inparticular those described in WO 2005/123101 and WO2008046676,skin-lightening agents, in particular those described in WO 2007/110415,skin-tanning agents, in particular those described in WO 2006/045760,cooling agents, in particular those described in WO 2005/123101,skin-cooling agents, in particular those described in WO 2005/123101,warming agents, in particular those described in WO 2005/123101,UV-absorbing agents, in particular those described in WO 2005/123101, UVfilters, in particular those described in WO 2005/123101,benzylidene-beta-dicarbonyl compounds in accordance with WO 2005/107692and alpha-benzoyl-cinnamic acid nitriles in accordance with WO2006/015954, insect repellents, in particular those described in WO2005/123101, plant parts, plant extracts, in particular those describedin WO 2005/123101, vitamins, in particular those described in WO2005/123101, emulsifiers, in particular those described in WO2005/123101, gelling agents, in particular those described in WO2005/123101, oils in particular those described in WO 2005/123101, waxesin particular those described in WO 2005/123101, fats in particularthose described in WO 2005/123101, phospholipids, in particular thosedescribed in WO 2005/123101, saturated fatty acids and mono- orpolyunsaturated fatty acids and α-hydroxy acids and polyhydroxy-fattyacids and esters of saturated and/or unsaturated branched and/orunbranched alkane carboxylic acids, in particular those described in WO2005/123101, surface-active substances (surfactants) in particular thosedescribed in WO 2005/123101, skin repair agents comprising cholesteroland/or fatty acids and/or ceramides and/or pseudoceramides, inparticular those described in WO 2006/053912, dyestuffs and colorantsand pigments, in particular those described in WO 2005/123101, aromachemicals and flavors and fragrances, in particular those described inS. Arctander, Perfume and Flavor Chemicals, private publishing house,Montclair, N J., 1969 and Surburg, Panten, Common Fragrance and FlavorMaterials, 5th Edition, Wiley-VCH, Weinheim 2006, preferably thoseexplicitly mentioned in US 2008/0070825, alcohols and polyols, inparticular those described in WO 2005/123101, organic solvents, inparticular those described in WO 2005/123101, silicones and siliconeoils and silicone derivatives in particular those described inWO2008046676, virucides, abrasives, anti-cellulite agents, astringents,antiseptic agents, antistatics, binders, buffers, cell stimulants,cleansing agents, care agents, depilatory agents, softeners, enzymes,essential oils, in particular those described in US 2008/0070825,fibres, film-forming agents, fixatives, foam-forming agents, foamstabilizers, substances for preventing foaming, foam boosters,gel-forming agents, hair growth activators, hair growth inhibitors, haircare agents, hair-setting agents, hair-straightening agents,hair-smoothening, bleaching agents, strengthening agents, stain-removingagents, optically brightening agents, impregnating agents,dirt-repellent agents, friction-reducing agents, lubricants, opacifyingagents, plasticizing agents, covering agents, polish, gloss agents,polymers in particular those described in WO2008046676, powders,peptides, mono-, di- and oligosaccharides, re-oiling agents, abradingagents, skin-soothing agents, skin-cleansing agents, skin care agents,skin-healing agents, skin-protecting agents, skin-softening agents,skin-smoothing agents, nourishing agents, skin-warming agents,stabilizers, detergents, fabric conditioning agents, suspending agents,thickeners, yeast extracts, algae or microalgae extracts, animalextracts, liquefiers, color-protecting agents, anticorrosives andelectrolytes.

Auxiliary substances and additives can be included in quantities of 5 to99.99 wt. %, preferably 10 to 80 wt. %, based on the total weight of theproduct. The amounts of cosmetic or dermatological auxiliary agents andadditives and perfume to be used in each case can easily be determinedby the person skilled in the art by simple trial and error, depending onthe nature of the particular product.

The products can also contain water in a quantity of up to 99.99 wt. %,preferably 5 to 80 wt. %, based on the total weight of the product.

Products according to the invention can contain one or more further hairgrowth modulating agents. A more rapid hair growth modulation based inpart on synergistic effects can be achieved in this way.

Agents to stimulate hair growth are for example pyrimidine derivativessuch as 2,4-diaminopyrimidine-3-oxide (Aminexil),2,4-diamino-6-piperidinopyrimidine-3-oxide (Minoxidil) and derivativesthereof, 6-amino-1,2-dihydro-1-hydroxy-2-imino-4-piperidinopyrimidineand its derivatives, pyridoxine hydrochloride, xanthine alkaloids suchas caffeine, theobromine and theophylline and derivatives thereof,6-(benzylamino)purin also known as cytokinin B, pantotenic acid and itsderivatives, quercetin and derivatives, dihydroquercetin (taxifolin) andderivatives, potassium channel openers, antiandrogenic agents, syntheticor natural 5-reductase inhibitors, nicotinic acid esters such astocopheryl nicotinate, benzyl nicotinate and C1-C6 alkyl nicotinate,proteins such as for example the tripeptide Lys-Pro-Val, diphencypren,hormons, finasteride, dutasteride, flutamide, bicalutamide, pregnanederivatives, progesterone and its derivatives, cyproterone acetate,spironolactone and other diuretics, calcineurin inhibitors such as FK506(Tacrolimus, Fujimycin) and its derivatives, Cyclosporin A andderivatives thereof, glyceryl monopentadecanoate or other promoters ofATP production in hair follicles, mononitro guaiacol, biotin, carproniumchloride, tocopherol and its derivatives, cepharanthine, sulphur,vitamin B6, glycyrrhetinic acid and its derivatives, inositol,hinokitiol, methionine, serine, threonine, zinc and zinc salts,polyphenols, procyanidins, proanthocyanidins, phytosterols such as forexample beta-sitosterol, biotin, eugenol, (±)-beta-citronellol,panthenol, glycogen for example from mussels, kankohso, placentalextract, royal yelly extract, Duku extract, extracts frommicroorganisms, algae, microalgae or plants and plant parts of forexample the genera dandelion (Leontodon or Taraxacum), Orthosiphon,Vitex, Coffea, Paullinia, Theobroma, Asiasarum, Cucurbita, Swertia,Capsicum or Styphnolobium, Serenoa repens (saw palmetto), Sophoraflavescens, Pygeum africanum, Panicum miliaceum, Cimicifuga racemosa,Glycine max, Eugenia caryophyllata, Excrementum bombycis, Cotinuscoggygria, Hibiscus rosa-sinensis, Camellia sinensis, Ilexparaguariensis, Polygonum multiflorum, licorice, grape, ginseng, ginkgo,apple, barley or hops or/nd hydrolysates from rice or wheat.

Agents to inhibit hair growth are for example activin, activinderivatives or activin agonists, ornithine decarboxylase inhibitors suchas alpha-difluoromethylomithine or pentacyclic triterpenes like forexample ursolic acid, betulin, betulinic acid, oleanolic acid andderivatives thereof, ornithine amino transferase inhibitors, serineproteases, 5alpha-reductase inhibitors, 5-lipoxygenase inhibitors,cyclooxygenase inhibitors, protein-tyrosine kinase inhibitors, proteinkinase C inhibitors, sulfotransferase inhibitors, nitric oxidesynthetase inhibitors, alkaline phosphatase inhibitors, inhibitors ofelastase-like enzymes, neutral endopeptidase inhibitors, matrixmetalloproteinase inhibitors, inhibitors of a cysteine pathway enzyme,androgen receptor antagonists, S-adenosylmethionine decarboxylaseinhibitors, gamma-glutamyl transpeptidase inhibitors, transglutaminaseinhibitors, VEGF modulators, compounds which block the glucose transferacross the membranes of the cells of hair follicles such as phloretin,soybean-derived serine protease inhibitors, extracts frommicroorganisms, algae, microalgae or plants and plant parts of forexample the families Leguminosae, Solanaceae, Graminae, Asclepiadaceaeor Cucurbitaceae, the genera Chondrus, Gloiopeltis, Ceramium, Durvillea,Glycine max, Sanguisorba officinalis, Calendula officinalis, Hamamelisvirginiana, Arnica montane, Salix alba, Hypericum perforatum or Gymnemasylvestre.

The amount of the aforementioned examples of additional activeingredients for the modulation of hair growth (one or more compounds) inthe formulations according to the invention is then preferably 0.00001to 30 wt. %, preferably 0.0001 to 20 wt. %, particularly preferably0.001 to 5 wt. %, based on the total weight of the product.

The products according to the invention can preferably also containother active ingredients which modulate skin and/or hair pigmentationand which are suitable for cosmetic (e.g. dermatological) and/ortherapeutic applications. A more rapid modulation of skin and/or hairpigmentation based in part on synergistic effects can be achieved inthis way.

Advantageous skin and hair lightening active ingredients in this respectare kojic acid (5-hydroxy-2-hydroxymethyl-4-pyranone), kojic acidderivatives e.g. kojic acid dipalmitate, arbutin, ascorbic acid,ascorbic acid derivatives, hydroquinone, hydroquinone derivatives,resorcinol, sulfur-containing molecules such as e.g. glutathione orcysteine, alpha-hydroxy acids (e.g. citric acid, lactic acid, malicacid) and derivatives thereof, N-acetyl tyrosine and derivatives,undecenoyl phenylalanine, gluconic acid, 4-alkyl resorcinols,4-(1-phenylethyl)-1,3-dihydroxybenzene, chromone derivatives such asaloesin, flavonoids, thymol derivatives, 1-aminoethyl phosphinic acid,thiourea derivatives, ellagic acid, nicotinamide (niacinamide), zincsalts such as e.g. zinc chloride or gluconate, thujaplicin andderivatives, triterpenes such as maslinic acid, sterols such asergosterol, benzofuranones such as senkyunolide, vinyl and ethylguiacol, dionic acids such as octodecene dionic acid and azelaic acid,inhibitors of nitrogen oxide synthesis, such as e.g. L-nitroarginine andderivatives thereof, 2,7-dinitroindazole or thiocitrulline, metalchelators (e.g. alpha-hydroxy fatty acids, palmitic acid, phytic acid,lactoferrin, humic acid, bile acid, bile extracts, bilirubin,biliverdin, EDTA, EGTA and derivatives thereof), retinoids, soya milkand extract, serine protease inhibitors or lipoic acid or othersynthetic or natural active ingredients for skin and hair lightening,the latter also being used in the form of an extract from plants, suchas e.g. bearberry extract, rice extract, papaya extract, liquorice rootextract or constituents concentrated therefrom, such as glabridin orlicochalcone A, artocarpus extract, extract of rumex and ramulusspecies, extracts of pine species (pinus) and extracts of vitis speciesor stilbene derivatives concentrated therefrom, extract of saxifrage,mulberry, scutelleria or/and grapes.

Advantageous skin and hair tanning active ingredients in this respectare substrates or substrate analogues of tyrosinase such as L-tyrosine,N-acetyl tyrosine, L-DOPA or L-dihydroxyphenylalanine, xanthinealkaloids such as caffeine, theobromine and theophylline and derivativesthereof, proopiomelanocortin peptides such as ACTH, alpha-MSH, peptideanalogues thereof and other substances which bind to the melanocortinreceptor, peptides such as Val-Gly-Val-Ala-Pro-Gly, Lys-Ile-Gly-Arg-Lysor Leu-Ile-Gly-Lys, purines, pyrimidines, folic acid, copper salts suchas copper gluconate, chloride or pyrrolidonate,1,3,4-oxadiazole-2-thiols such as5-pyrazin-2-yl-1,3,4-oxadiazole-2-thiol, curcumin, zinc diglycinate(Zn(Gly)2), manganese(II) bicarbonate complexes (“pseudocatalases”) asdescribed for example in EP 0 584 178, tetrasubstituted cyclohexenederivatives as described for example in WO 2005/032501 isoprenoids asdescribed in WO 2005/102252 and in WO 2006/010661, melanin derivativessuch as Melasyn-100 and MelanZe, diacyl glycerols, aliphatic or cyclicdiols, psoralens, prostaglandins and analogues thereof, activators ofadenylate cyclase and compounds which activate the transfer ofmelanosomes to keratinocytes such as serine proteases or agonists of thePAR-2 receptor, extracts of plants and plant parts of the chrysanthemumspecies, sanguisorba species, walnut extracts, urucum extracts, rhubarbextracts, trehalose, erythrulose and dihydroxyacetone. Flavonoids whichbring about skin and hair tinting or browning (e.g. quercetin,rhamnetin, kaempferol, fisetin, genistein, daidzein, chrysin andapigenin, epicatechin, diosmin and diosmetin, morin, quercitrin,naringenin, hesperidin, phloridzin and phloretin) can also be used.

The amount of the aforementioned examples of additional activeingredients for the modulation of skin and hair pigmentation (one ormore compounds) in the products according to the invention is thenpreferably 0.00001 to 30 wt. %, preferably 0.0001 to 20 wt. %,particularly preferably 0.001 to 5 wt. %, based on the total weight ofthe preparation.

Products according to the present invention in the form of cosmeticand/or dermatologically active products are applied to the skin and/orhair in a sufficient amount in the conventional manner for cosmetics anddermatics. In this context, cosmetic and dermatological productsaccording to the present invention which additionally act as sunscreenproducts offer particular advantages. These products (formulations)advantageously comprise at least one UVA filter and/or at least one UVBfilter and/or at least one inorganic pigment. In this context, theproducts can be in various forms such as are conventionally employede.g. for sunscreen formulations. They can be e.g. a solution, anemulsion of the water-in-oil (W/O) type or of the oil-in-water (O/W)type or a multiple emulsion, for example of the water-in-oil-in-water(W/O/W) type, a gel, a hydrodispersion, a solid stick or also anaerosol.

As mentioned, products according to the present invention canadvantageously be combined with substances which absorb UV radiation,the total amount of the filter substances being e.g. 0.01 to 40 wt.-%,preferably 0.1 to 10 wt.-%, in particular 1.0 to 5.0 wt.-%, based on thetotal weight of the formulations, in order to provide cosmetic productswhich protect the hair or skin from ultraviolet radiation.

Preferred products of the present invention are sunscreen formulationsin the form of aqueous emulsions, preferably of the water-in-oil (W/O)or of the oil-in-water (O/W) type or a multiple emulsion, for example ofthe water-in-oil-in-water (W/O/W) type, more preferably of theoil-in-water (O/W) type.

Preferred sunscreen formulations (products) of the present inventioncomprise a total amount of organic UV filters of greater than 10 wt.-%,preferably in the range of from 12 to 40 wt.-%, more preferred in therange of from 15 to 35 wt.-%, based on the total weight of the sunscreenformulation.

In this context advantageous organic UV filters are:

-   -   p-aminobenzoic acid    -   p-aminobenzoic acid ethyl ester (25 mol) ethoxylated    -   p-dimethylaminobenzoic acid-2-ethylhexyl ester    -   p-aminobenzoic acid ethyl ester (2 mol)N-propoxylated    -   p-aminobenzoic acid glycerol ester    -   salicylic acid homomenthyl ester (homosalates) (Neo        Heliopan®HMS)    -   salicylic acid-2-ethylhexyl ester (Neo Heliopan®OS)    -   triethanolamine salicylate    -   4-isopropyl benzyl salicylate    -   anthranilic acid menthyl ester (Neo Heliopan®MA)    -   diisopropyl cinnamic acid ethyl ester    -   p-methoxycinnamic acid-2-ethylhexyl ester (Neo Heliopan®AV)    -   diisopropyl cinnamic acid methyl ester    -   p-methoxycinnamic acid isoamyl ester (Neo Heliopan®E 1000)    -   p-methoxycinnamic acid diethanolamine salt    -   p-methoxycinnamic acid isopropyl ester    -   2-ethylhexyl-2-cyano-3,3-diphenyl acrylate (Neo Heliopan®303)    -   ethyl-2-cyano-3,3′-diphenyl acrylate    -   2-phenylbenzimidazole sulfonic acid and salts (Neo        Heliopan®Hydro)    -   3-(4′-trimethylammonium) benzylidene bornan-2-one methyl sulfate    -   terephthalylidene dibornane sulfonic acid and salts (Mexoryl®SX)    -   4-t-butyl-4′-methoxydibenzoyl methane (avobenzone)/(Neo        Heliopan®357)    -   β-imidazole-4(5)-acrylic acid (urocanic acid)    -   2-hydroxy-4-methoxybenzophenone (Neo Heliopan®BB)    -   2-hydroxy-4-methoxybenzophenone-5-sulfonic acid    -   dihydroxy-4-methoxybenzophenone    -   2,4-dihydroxybenzophenone    -   tetrahydroxybenzophenone    -   2,2′-dihydroxy-4,4′-dimethoxybenzophenone    -   2-hydroxy-4-n-octoxybenzophenone    -   2-hydroxy-4-methoxy-4′-methyl benzophenone    -   3-(4′-sulfo)benzylidene bornan-2-one and salts    -   3-(4′-methyl benzylidene)-d,l-camphor (Neo Heliopan®MBC)    -   3-benzylidene-d,l-camphor    -   4-isopropyl dibenzoyl methane    -   2,4,6-trianilino-(p-carbo-2′-ethylhexyl-1′-oxy)-1,3,5-triazine    -   phenylene bis-benzimidazyl tetrasulfonic acid disodium salt (Neo        Heliopan®AP)    -   2,2′-(1,4-phenyleneybis-(1H-benzimidazole-4,6-disulfonic acid),        monosodium salt    -   N-[(2 and 4)-[2-(oxoborn-3-ylidene) ethyl]benzyl] acrylamide        polymer    -   phenol,        -(2H-benzotriazol-2-yl)-4-methyl-6-(2-methyl-3(1,3,3,3-tetramethyl-1-(trimethylsilyl)oxy)disiloxyanyl)        propyl), (Mexory®XL)    -   4,4′-[(6-[4-(1,1-dimethyl)aminocarbonyl)        phenylamino]-1,3,5-triazine-2,4-diyl)diimino]-bis-(benzoic        acid-2-ethylhexyl ester) (Uvasorb®HEB)    -   2,2′-methylene        bis-(6-(2H-benzotriazol-2-yl)-4-1,1,3,3-tetramethylbutyl)        phenol), (Tinosorb®M)    -   2,4-bis-[4-(2-ethylhexyloxy)-2-hydroxyphenyl]-1,3,5-triazine    -   benzylidene malonate polysiloxane (Parsol®SLX)    -   glyceryl ethylhexanoate dimethoxycinnamate    -   disodium-2,2′-dihydroxy-4,4′-dimethoxy-5,5-disulfobenzophenone    -   dipropylene glycol salicylate    -   sodium hydroxymethoxybenzophenone sulfonate    -   4,4′,4-(1,3,5-triazine-2,4,6-triyltriimino)-tris-benzoic acid        tris(2-ethylhexyl ester) (Uvinul®T150)    -   2,4-bis-[{(4-(2-ethylhexyloxy)-2-hydroxy}phenyl]-6-(4-methoxyphenyl)-1,3,5-triazine        (Tinosorb®S)    -   2,4-bis-[{(4-(3-sulfonato)-2-hydroxypropyloxy)-2-hydroxy}phenyl]-6-(4-methoxyphenyl)-1,3,5-triazine        sodium salt    -   2,4-bis-[{(3-(2-propyloxy)-2-hydroxypropyloxy)-2-hydroxy}phenyl]-6-(4-methoxyphenyl)-1,3,5-triazine    -   2,4-bis-[{4-(2-ethylhexyloxy)-2-hydroxy}phenyl]-6-[4-(2-methoxyethyl        carbonyl)phenylamino]-1,3,5-triazine    -   2,4-bis-[{4-(3-(2-propyloxy)-2-hydroxypropyloxy)-2-hydroxy}phenyl]-6-[4-(2-ethylcarboxyl)        phenylamino]-1,3,5-triazine    -   2,4-bis-[{4-(2-ethylhexyloxy)-2-hydroxy}phenyl]-6-(1-methylpyrrol-2-yl)-1,3,5-triazine    -   2,4-bis-[{4-tris-(trimethylsiloxysilylpropyloxy)-2-hydroxy}phenyl]-6-(4-methoxyphenyl)-1,3,5-triazine    -   2,4-bis-[{4-(2″-methylpropenyloxy)-2-hydroxy}phenyl]-6-(4-methoxyphenyl)-1,3,5-triazine    -   2,4-bis-[{4-(1′,1′,1′,3′5′,5′,5′-heptamethylsiloxy-2″-methylpropyloxy)-2-hydroxy}phenyl]-6-(4-methoxyphenyl)-1,3,5-triazine    -   2-(4-diethylamino-2-hydroxybenzoyl) benzoic acid hexyl ester        (Uvinul® A Plus)    -   indanylidene compounds in accordance with WO 02/38537

Organic UV filters which are particularly preferred in products of thepresent invention (in particular if they are in the form of a sunscreenformulation), preferably in an amount mentioned (above), are:

-   -   p-aminobenzoic acid    -   3-(4′-trimethylammonium) benzylidene bornan-2-one methyl sulfate    -   salicylic acid homomenthyl ester (Neo Heliopan®HMS)    -   2-hydroxy-4-methoxybenzophenone (Neo Heliopan®BB)    -   2-phenylbenzimidazole sulfonic acid (Neo Heliopan®Hydro)    -   terephthalylidene dibornane sulfonic acid and salts (Mexoryl®SX)    -   4-tert-butyl-4′-methoxydibenzoyl methane (Neo Heliopan®357)    -   3-(4′-sulfo)benzylidene bornan-2-one and salts    -   2-ethylhexyl-2-cyano-3,3-diphenyl acrylate (Neo Heliopan®303)    -   N-[(2 and 4)-[2-(oxoborn-3-ylidene) methyl]benzyl] acrylamide        polymer    -   p-methoxycinnamic acid-2-ethylhexyl ester (Neo Heliopan®AV)    -   p-aminobenzoic acid ethyl ester (25 mol) ethoxylated    -   p-methoxycinnamic acid isoamyl ester (Neo Heliopan®E1000)    -   2,4,6-trianilino-(p-carbo-2′-ethylhexyl-1′-oxy)-1,3,5-triazine        (Uvinul®T150)    -   phenol,        2-(2H-benzotriazol-2-yl)-4-methyl-6-(2-methyl-3(1,3,3,3-tetramethyl-1-(trimethylsilyl)oxy)disiloxyanyl)        propyl), (Mexoryl®XL)    -   4,4′4-[(6-[4-(1,1-dimethyl)aminocarbonyl)        phenylamino]-1,3,5-triazine-2,4-diyl)diimino]-bis-(benzoic        acid-2-ethylhexyl ester) (Uvasorb HEB)    -   3-(4′-methyl benzylidene)-d,l-camphor (Neo Heliopan®MBC)    -   3-benzylidene camphor    -   salicylic acid-2-ethylhexyl ester (Neo Heliopan®OS)    -   4-dimethylaminobenzoic acid-2-ethylhexyl ester (Padimate O)    -   hydroxy-4-methoxybenzophenone-5-sulfonic acid and Na salt    -   2,2′-methylene        bis-(6-(2H-benzotriazol-2-yl)-4-1,1,3,3-tetramethylbutyl)        phenol) (Tinosorb®M)    -   phenylene bis-benzimidazyl tetrasulfonic acid disodium salt (Neo        Heliopan®AP)    -   2,4-bis-[{(4-(2-ethylhexyloxy)-2-hydroxy}phenyl]-6-(4-methoxyphenyl)-1,3,5-triazine,        (Tinosorb®S)    -   benzylidene malonate polysiloxane (Parsol®SLX)    -   menthyl anthranilate (Neo Heliopan®MA)    -   2-(4-diethylamino-2-hydroxybenzoyl) benzoic acid hexyl ester        (Uvinul® A Plus)    -   indanylidene compounds in accordance with WO 02/38537

Products according to the present invention in the form of sunscreenformulations preferably have a SPF (sun protection factor) of equal orgreater than 15, preferably of equal or greater than 20, more preferablyof equal or greater than 30.

Preferred products of the present invention in the form of sunscreenformulations comprise 4-(1,1-dimethylethyl)-4′-methoxydibenzoylmethane(4-t-butyl-4′-methoxydibenzoyl methane; avobenzone), preferably in anamount in the range of from 0.2-10 wt.-%, more preferred in the range offrom 0.5-5 wt.-%, based on the total weight of the sunscreenformulation.

In preferred sunscreen formulations comprising components (a) and (b)the pH-value is in the range of from pH 4 to pH 8, preferably from pH 4to 6.5.

Products according to the present invention may comprise one or morecompatible solutes. Preferred compatible solutes are described in WO01/76572, namely dimyo-inositol phosphate (DIP), diglycerin phospate(DGP), di-myo-inositol phosphate (DIP), cyclic 2,3 diphosphoglycerate(cDPG), 1,1-di-glycerol phosphate (DGP), beta-mannosyl glycerate(firoin), beta-mannosyl glyceramide (firoin-A) anddi-mannosyl-di-inositol phosphate (DMIP) and ectoine andectoine-derivatives, as described in EP 0 553 884 A, EP 0 671 161 A andWO 94/15923, in particular((S)-1,4,5,6-tetrahydro-2-methyl-4-pyrimidinecarboxylic acid) andhydroxyectoine((S,S)-1,4,5,6-tetrahydro-5-hydroxy-2-methyl-4-pyrimidinecarboxylicacid).

Preferably, the total amount of compatible solutes is in the range offrom 0.05 to 10 wt.-%, preferably 0.1 to 5 wt.-%, based on the totalweight of the product according to the present invention.

Also preferred are products according to the present inventioncomprising one or more cooling agents selected from the group consistingof: menthol, preferably l-menthol, menthone glycerin acetal (trade name:Frescolat®MGA), menthyl lactate (preferably 1-menthyl lactate, inparticular l-menthyl-l-lactate, trade name: Frescolat®ML), substitutedmenthyl-3-carboxylic acid amide (e.g. menthyl-3-carboxylicacid-N-ethylamide), 2-isopropyl-N-2,3-trimethylbutanamide, substitutedcyclohexane carboxylic acid amide, 3-menthoxypropane-1,2-diol,2-hydroxyethylmenthylcarbonate, 2-hydroxypropylmenthylcarbonate,N-acetyl glycine menthyl ester, Isopulegol, menthyl hydroxycarboxylicacid ester (e.g. menthyl-3-hydroxybutyrate), monomenthylsuccinate,2-mercaptocyclodecanone, menthyl-2-pyrrolidin-5-one carboxylate,2,3-dihydroxy-p-menthane, 3,3,5-trimethyl cyclohexanone glycerine ketal,3-menthyl-3,6-di- and -trioxa Ikanoate, 3-menthylmethoxy acetate,N-(4-cyanomethylphenyl)-p-menthanecarboxamide,N-(4-cyanophenyl)-p-menthanecarboxamide and Icilin.

Preferred cooling agents are: 1-menthol, menthone glycerine acetal(trade name: Frescolat®MGA), menthyl lactate (preferably l-menthyllactate, in particular l-menthyl-l-lactate, trade name: Frescolat®ML),3-menthoxy propane-1,2-diol, 2-hydroxyethyl menthyl carbonate,2-hydroxypropyl menthyl carbonate, particular preference being forl-menthyl-l-lactate.

Products according to the present invention may comprise one or moreanti-cellulite agents as well as agents enhancing or boosting theactivity of anti-cellulite agents.

Anti-cellulite agents and lipolytic agents are preferably selected fromthe group consisting of those described in WO 2007/077541, andbeta-adrenergic receptor agonists such as synephrine and itsderivatives.

Agents enhancing or boosting the activity of anti-cellulite agents, inparticular agents which stimulate and/or depolarise C nerve fibres, arepreferably selected from the group consisting of capsaicin andderivatives thereof, vanillyl-nonylamid and derivatives thereof,L-carnitine, coenzym A, isoflavonoides, soy extracts, ananas extract andconjugated linoleic acid.

For certain products according to the present invention are preferredwhich in addition comprise one, two or more compounds of the groupconsisting of:

glycerol, 1,2-propanediol, 1,3-propanediol, 1,2-butanediol,1,3-butanediol, 1,4-butanediol, 2,3-butanediol, 1,2-pentanediol,1,5-pentanedial, 1,2-hexanediol, 1,6-hexanediol, 1,2-octanediol,1,8-octanediol, 2-methylpentane-2,4-diol, 2,5-hexanediol,3,6-octanediol, 2-ethyl-1,3-hexanediol, 1,3-octanediol, 1,2-decanediol,1,3-decanediol, 1,2-dodecandiol, 1,2-tetradecandiol.

Products of the present invention in addition may additionally compriseknown antimicrobials like chitosan, totarol, farnesol, glycerolmonolaurate, arylalkyl alcohols, such as e.g.4-methyl-4-phenyl-2-pentanol and its derivatives (DE 101 43 434, inparticular 4-methyl-4-phenyl-2-pentanol), muguet alcohol(2,2-dimethyl-3-phenylpropanol), other arylalkyl alcohols (e.g. asdisclosed in DE 44 47 361, DE 103 30 697, U.S. Pat. No. 4,110,430 or EP1 157 687), 2-butyloctanoic acid, 2-hexyldecanoic acid, p-anisic acid,essential oils with antimicrobial properties and isolates from essentialoils with antimicrobial properties like e.g. thymol or eugenol, perfumeoils or single aroma chemicals with antimicrobial activity, polyglycerolesters, such as e.g. polyglyceryl 3-caprylates, or combinations of thesubstances mentioned, which are generally employed, inter alia, againstunderarm odor, foot odor, acne or dandruff formation.

In products of the present invention a combination with (metal)chelators is advantageous in some cases. (Metal) chelators which arepreferably to be employed here are, inter alia, α-hydroxy fatty acids,phytic acid, lactoferrin, α-hydroxy acids, such as, inter alia, citricacid, lactic acid and malic acid, and humic acids, bile acids, bileextracts, bilirubin, biliverdin or EDTA, EGTA and derivatives thereof.

BRIEF DESCRIPTION OF DRAWINGS AND FIGURES

FIG. 1 shows the increase of hair follicles at day 10 of culture,expressed as percentage relative to an untreated control (modulation bydir-MeOH).

FIG. 2 shows the increase of hair follicles at day 10 of culture,expressed as percentage relative to an untreated control (modulation bydir-EtOH).

FIG. 3 shows the increase of hair follicles at day 10 of culture,expressed as percentage relative to an untreated control (modulation bydir-EtAc).

FIG. 4 shows the increase of hair follicles at day 10 of culture,expressed as percentage relative to an untreated control (modulation bysequential extracts).

FIG. 5 shows the increase of hair follicles at day 10 of culture,expressed as percentage relative to an untreated control (modulation bydir-Hex).

FIG. 6 shows the increase of hair follicle melanin content at day 10 ofculture, expressed as percentage relative to an untreated control(modulation by dir-EtAc and SEQ-3-% EtOH).

FIG. 7 shows the increase of hair follicle melanin content at day 10 ofculture, expressed as percentage relative to an untreated control(modulation by sequential extraction).

FIG. 8 shows the increase in skin pigmentation at day 10 of culture,expressed as percentage relative to an untreated control (modulation bysequential extracts).

FIG. 9 shows the increase in skin pigmentation at day 10 of culture,expressed as percentage relative to an untreated control (modulation bydir-EtOH).

FIG. 10 shows the increase in skin pigmentation at day 10 of culture,expressed as percentage relative to an untreated control (comparison ofdir-EtOH and sequential extracts).

The invention is further described by the following figures andexamples, without limiting the scope of the claims.

Extraction Example 1: Preparation of “Direct Extracts”

Tahitian Isochrysis CS 177 was used to prepare extracts by the followingsteps:

1. Prepare a suspension of powdery freeze-dried Isochrysis cell materialin the selected extractant at a ratio (dry weight/volume) of 10 mg:1 ml;2. stir the suspension in the dark at room temperature for 16 h;3. centrifuge the suspension at 2000 g for 15 minutes to recover asupernatant and a cell material pellet;4. resuspend the pellet in 0.5 ml of the aforementioned selectedextractant for each ml used at the step 1;5. immediately centrifuge the suspension at 2000 g for 15 minutes torecover a supernatant and a cell material pellet;6. repeat steps 4 and 5 one more time;7. combine the supernatants to form the “direct extract” of therespective extractant.

The extractant was chosen from water, methanol, ethanol, isopropanol,ethyl acetate and hexane:

TABLE 1 Ratio of dry extract/biomass (dry weight) with differentextractants % of the Extractant dried biomass Water 54 Methanol 47Ethanol 38 Iso-propanol 24 Ethyl acetate 20 Hexane 12

Extraction Example 2: Preparation of “Sequential Extracts”

For preparation of “sequential extracts”, first a direct extract wasprepared. After step 6, the cell material pellet was resuspended in aselected further extractant, and steps 3 to 7 were then performed againwith the selected further extractants.

A series of sequential extractions was produced according to table 2:

TABLE 2 sequential extracts and related ratio of extract/biomass in dryweight % ratio dry Sequential Extract extract/ Version protocolExtractant symbol biomass 1 Ethyl acetate Ethyl acetate dir-EtAc 20followed by 30% ethanol seq. 30% 30% ethanol EtOH 2 Methanol methanoldirMeOH 47 followed by water water seq water2 15 3 Hexane Hexane dir-Hex12 followed by ethyl ethyl acetate seq-EtAc 6 acetate followed byethanol Ethanol Seq-EtOH 17 followed by water Water Seq-Water 31 4 Ethylacetate Ethyl acetate dir-EtAc 20 followed by ethanol Ethanol seq-EtOH218 followed by water Water seq-Water3 30 5 Methanol methanol dir-MeOH 47followed by hexane/ hexane/ethyl seq- 3 ethyl acetate 1:1 acetate 1:1Hex/EtAc (v/v) (v/v) followed by water water Seq-Water4 7

Effects Examples 1-3: Modulation of Hair Follicle Growth by Dir-MeOH

These examples demonstrate the influence of the “direct methanolextract” according to extraction example 1 on hair follicle metabolism.

For each experiment and the control 9-12 follicles were used, plated atthe density of 3 hair follicles/well in 24 well plates. Hair follicleswere taken from the head of the scalp and transferred for cultivation insterile 24 well plates using a modified Williams' Medium E. Cultivationtook place for nine days, following 18 h of pre-incubation performed inorder to select hair follicles suitable to be maintained in culture.Only those follicles showing a good vital stage and a growth of not lessthan 0.2 mm were used.

The growth performances observed in the treated hair follicles werecompared to a control group, which was cultured in the same culturemedium but free from extract supplement.

The experimental design consisted in treatments with dir-MeOH extract atthree final concentrations corresponding to 0.1, 1 and 10 μg/ml; theconcentrations were calculated in terms of extracted freeze-driedbiomass. In order to obtain these supplemented media, the requiredquantity of dir-MeOH extract was submitted to solvent evaporation andthen redissolved again in DMSO. The final concentration of thisDMSO-solved extract has been adjusted in order to supplement theexperimental media with the desired extract quantity, obtaining at sametime a final concentration in DMSO equal to 0.05%. The sameconcentration of DMSO has also been used in the medium for the cultureof the control group.

The activity of the microalgae treatment is demonstrated by the increaseof hair follicle growth expressed as percentage variation in comparisonto the elongation performed by the control group. The experiments wereterminated after 10 days of cultivation (9 days of treatment). Thegrowth of the hair follicles was studied by microphotography andsubsequently determined by image analysis. All the hair follicles werephotographed every two days.

The described experiment was replicated three times adopting hairfollicles taken from 3 different donors. The results were pooled andcombined into table 3 and FIG. 1, were the hair follicle elongation isexpressed as percentage ratio between the experimental groups and theuntreated control:

TABLE 3 Growth of hair follicles at day 10 of culture-Data pooled fromthree replicates Elongation in [%] of the control performance ± standarderror Experi- No. DHA PUFAs mental Stan- of medium medium treat- Growthdard Hair ANOVA content content Ex. ment (%) error follicles Test wt. %wt. % 0 Control 100 3.8 28 — — 1 dir- 118.1 3.9 25 P < 0.01 0 0.01 MeOHextr. 0.1 μg/ml 2 dir- 109.6 3.3 24 n.s. 0.02 0.06 MeOH extr. 1 μg/ml 3dir- 108.7 4.8 24 n.s. 0.24 0.56 MeOH extr. 10 μg/ml DHA:docosahexaenoic acid, C22:6, the most prominent polyunsaturated fattyacid (PUFA) PUFAs: DHA (C22:6), EPA (C20:5), stearidonic acid (C18:4),linolenic (C18:3) and linolic acid (C18:2)

The results are also shown in FIG. 1.

The results indicate that the addition of the dir-MeOH extract leads toa significant increase in growth of the hair follicles, varying from 9to 18% in comparison to the untreated group. The most significantresponse has been obtained at the lower treatment which results highlysignificant also on a statistical basis (P<0.01).

Effects Examples 4-5: Modulation of Hair Growth by Dir-EtOH

The same experimental protocol described for the examples 1-3 has beenrepeated to investigate the activity of the direct ethanol extract withthe exception that all the experimental groups and the control wereprepared comprising 12-18 follicles.

The following data has been obtained treating hair follicles taken fromthree different donors. Table 4 and FIG. 2 summarize the results.

TABLE 4 Growth of hair follicles at day 10 of culture-Data pooled fromthree replicates Elongation in [%] of the control performance ± standarderror PU- FAs * DHA medium Experi- Stan- medium content mental Growthdard N. of Hair ANOVA content wt. Ex. treatment (%) error follicles Testwt. % % 0 Control 100 2.4 44 — — 4 dr-EtOH 91.9 4.2 30 0 0.01 extr. 0.1μg/ml 5 dir-EtOH 81.2 2.4 31 P < 0.01 0.28 0.87 extr. 10 μg/ml * DHA(C22:6), EPA (C20:5), stearidonic acid (C18:4); linolenic (C18:3) andlinolic acid (C18:2)

The results indicate that the addition of the EtOH extract leads to asignificant reduction in growth of the hair follicles, varying from 8 to19% in comparison to the untreated group. The most significant responsehas been obtained at the 10 μg/ml treatment, which results highlysignificant also on a statistical basis (P<0.01).

Effects Examples 6-8: Modulation of Hair Growth by Dir-EtAc

The same experimental protocol described for the effects examples 1-3was repeated to investigate the activity of the direct ethyl acetateextract.

Table 5 and FIG. 3 summarize the results obtained by three replicateexperiments using different donors:

TABLE 5 Growth of hair follicles at day 10 of culture-Data pooled fromthree replicates Elongation in [%] of the control performance ± standarderror Experi- No. DHA PUFAs* mental Stan- of medium medium treat- Growthdard Hair ANOVA content content Ex. ment (%) error follicles Test wt. %wt. % 0 Control 100 3.8 28 — — 6 Dir-EtAc 95.3 4.6 26 n.s. 0 0.01 extr.0.1 μg/ml 7 Dir-EtAc 112.2 44 25 P < 0.05 0.03 0.07 extr. 1 μg/ml 8Dir-EtAc 105.9 4.5 26 n.s. 0.31 0.7 extr. 10 μg/ml *DHA (C22:6), EPA(C20:5), stearidonic acid (C18:4), linolenic (C18:3) and linolic acid(C18:2)

The data show the effectiveness of the extract for modulation of thehair follicle growth. The treatment at middle concentration (1 μg/ml)produced a significant stimulation (P<0.05) of hair follicle growthequal to 12%, while increasing or reducing the treatment intensity themodulation become not significant.

Effects Examples 9-20: Modulation of Hair Growth by Sequential Extracts

“Sequential extracts” were prepared as described in extraction example2. Four extracts were prepared (dir-HEX followed by seq-EtAc followed byseq-EtOH followed by seq-Water) and three dose treatments were testedfor each of them. The experiment has been repeated three times usinghair follicles obtained from different donors and the data have beenpooled to represent the average response of the three donors. Table 6and FIG. 4 summarize the results:

TABLE 6 Growth of hair follicles at day 10 of culture-Data pooled fromthree replicates Elongation in [%] of the control performance ± standarderror Experi- No. DHA PUFAs* mental Stan- of medium medium treat- Growthdard Hair ANOVA content content Ex. ment (%) error follicles Test wt. %wt. % 0 Control 100 3.6 35 — — 9 Dir- 92.5 3.6 34 n.s. 0 0.01 Hexextract 0.1 μg/ml 10 Dir- 97.6 3.6 34 n.s. 0.03 0.06 Hex extract 1 μg/ml11 Dir- 108.0 4.0 34 n.s. 025 0.58 Hex extract 10 μg/ml 12 Seq- 98.3 3.634 n.s. 0 0 EtAc extract 0.1 μg/ml 13 Seq- 100.3 3.6 31 n.s. 0 0.01 EtAcextract 1 μg/ml 14 Seq- 98.9 3.5 30 n.s. 0.04 0.09 EtAc extract 10 μg/ml15 Seq- 99.2 3.6 32 n.s. 0 0 EtOH extract 0.1 μg/ml 16 Seq- 96 3.1 34n.s. 0 0 EtOH extract 1 μg/ml 17 Seq- 88.6 2.9 30 P < 0.05 0.02 0.03EtOH extract 10 μg/ml 18 Seq- 94.4 4.7 32 n.s. 0 0 Water extract 0.1μg/ml 19 Seq- 95.8 3.1 32 n.s. 0 0 Water extract 1 μg/ml 20 Seq- 99 5.328 n.s. 0 0 Water extract 10 μg/ml *DHA (C22:6), EPA (C20:5),stearidonic acid (C18:4), linolenic (C18:3) and linolic acid (C18:2)

The experiments show that two classes of compounds active on hair growthhave been separated in the sequential extracts: the first has beenextracted by hexane and it produced a modulation of the hair growthchanging from inhibiting to stimulating in response to the increasingintensity of treatment, while the second one is more hydrophilic and hasbeen extracted by seq-ethanol. This latter produced a significantinhibition of the hair follicle growth (P<0.05) at 10 μg/ml.

Effects Examples 21-26: Modulation of Hair Growth by Direct HexaneExtracts

Direct hexane extract were prepared as described in Extraction Example 1to investigate the activity of the direct extract obtained using hexanein comparison with the activity of pure DHA (docosahexaenoic acid,C22:6). The aim of the experiment was to verify that the action of thedir-Hex is reproducible by treatment with DHA that is highly representedamong the PUFAs synthesised by Isochrysis sp., in particular TahitianIsochrysis. Since it has been detected that the dir-Hex extract isconstituted of DHA for about 20%, it has been estimated that a treatmentwith dir-Hex at 0.1 μg/ml corresponds to treat hair follicles with 0.02μg/ml of DHA. In order to reproduce a treatment balanced around thisvalue three groups of hair follicles were cultured in medium culturesupplemented with DHA ranging from 0.003 to 0.3 μg/ml and they have beencompared with others treated with dir-Hex ranging from 0.1 to 10 μg/ml,as usual. In order to supplement the culture medium with DHA,docosahexaenoic acid was solved in DMSO at concentrations suitable toarrive at the final content of 0.05% DMSO in the medium. The experimenthas been repeated twice and the pooled results are summarised in Table 7and FIG. 5.

TABLE 7 Growth of hair follicles at day 10 of culture-Data pooled fromtwo donors Elongation in [%] of the control performance ± standard errorExperi- No. DHA PUFAs mental Stan- of medium medium treat- Growth dardHair ANOVA content content Ex. ment (%) error follicles Test wt. % wt. %0 Control 100 3.3 30 — — 21 DHA 99.5 3.6 19 0.003 μg/ml 22 DHA 93.5 4.718 0.03 μg/ml 23 DHA 89.3 3.5 19 P < 0.05 0.3 μg/ml 24 Dir- 88.4 3.2 19P < 0.05 0 0.01 Hex extract 0.1 μg/ml 25 Dir- 98 4.3 19 0.03 0.06 Hexextract 1 μg/ml 26 Dir- 110.1 4.8 19 0.25 0.58 Hex extract 10 μg/ml

The results show that DHA produced an inhibiting dose-response, whilethe dir-Hex inhibited hair follicle growth at low concentration (i.e.low DHA) and stimulated the growth at higher concentrations (i.e.increasing DHA content). These data confirm that DHA and dir-Hextreatments produce different effects on hair follicle growth and thedir-Hex activity cannot be explained by DHA content.

Effects Examples 27-30: Modulation of Hair Pigmentation by Dir-Etac andSeq-30% EtOH

The activity on pigmentation of both lipophilic (dir-EtAc) andhydrophilic (seq-30% EtOH) extracts obtained from Tahitian Isochrysishas been studied by performing an experiment with a biological sampletaken from a single donor. The culture media for the experimentaltreatments were prepared according to the previous Extraction Examples,in order to obtain the following experimental treatments:

1) dir-EtAc=0.1 μg/ml and 1 μg/ml;2) seq-30% EtOH=0.1 μg/ml and 10 μg/ml.

The hair follicles culture was terminated after 5 days of cultivation (4of treatment). Subsequently, hair follicles were subjected tohistological analysis by preparing sections stained according to FontanaMasson technique. The melanin content of the tissues surroundingdermopapillas was detected by image analysis and the results are shownin Table 8 and FIG. 6.

TABLE 8 Melanin content of hair follicles at day 5 of culture - Datafrom single donor Melanin content in [%] of the control performance ±standard error Experimental Melanin Standard N. of ANOVA Ex. Treatmentcontent (%) error Samples Test 0 Control 100 21.5 12 27 dir-EtAc 112.318.7 12 0.1 μg/ml 28 dir-EtAc 149.8 36.8 12 1 μg/ml 29 seq- 111.4 22.812 30% EtOH 0.1 μg/ml 30 seq- 223.5 45.7 12 P < 0.01 30% EtOH 10 μg/ml

The results clearly indicate that the treatment of hair follicles withthe microalgae extracts increased the content of melanin after 4 days oftreatment. The intensity of the response varies with the dose. However,the more intensive responses have been detected by treating with seq-30%EtOH extract at 10 μg/ml (highly significant; P<0.01) and dir-Et-Acextract at 1 μg/ml.

Effects Examples 31-42: Modulation of Hair Pigmentation by SequentialExtracts

The experiment on hair follicle pigmentation has been repeated usingfour-step sequential extracts (hexane followed by ethyl acetate followedby ethanol followed by water) prepared by treating the Isochrysisbiomass as previously described in Extraction Example 2.

The culture techniques and histological analysis were the same describedfor Effects Examples 27-30.

The culture media for the experimental treatments were preparedaccording to the previous descriptions, in order to obtain the followingexperimental treatments:

1) dir-Hexane=0.1-1-10 μg/ml;2) seq-EtAc=0.1-1-10 μg/ml;3) seq-EtOH=0.1-1-10 μg/ml;4) seq-water=0.1-1-10 μg/ml;

The results are shown in Table 9 and FIG. 7:

TABLE 9 Melanin content of hair follicles at day 5 of culture - Datafrom single donor Melanin content in [%] of the control performance ±standard error Experimental Melanin Standard N. of ANOVA Ex. Treatmentcontent (%) error Samples Test 0 Control 100 13.3 12 31 dir-Hex 93.714.5 12 0.1 μg/ml 32 dir-Hex 125 13.3 12 1 μg/ml 33 dir-Hex 114.8 13.812 10 μg/ml 34 seq-EtAc 115.4 17.8 12 0.1 μg/ml 35 seq-EtAc 113.6 18.912 1 μg/ml 36 seq-EtAc 153.3 6.8 12 P < 0.05 10 μg/ml 37 seq-EtOH 117.618.3 12 0.1 μg/ml 38 seq-EtOH 115.3 14.3 12 1 μg/ml 39 seq-EtOH 108.1 1612 10 μg/ml 40 seq-Water 136 14.6 12 0.1 μg/ml 41 seq-Water 112.8 9 12 1μg/ml 42 seq-Water 163.2 9.8 12 P < 0.01 10 μg/ml

The results point out a general pigmentation enhancement in response toexperimental treatments, with statistically significant results for 10μg/ml seq-EtAc (ANOVA=P<0.05) and 10 μg/ml seq-water (ANOVA=P<0.01).

Effects Examples 43-50: Modulation of Skin Pigmentation by SequentialExtracts

Organ culture of full thickness human skin has been performed startingfrom a skin sample, exciding pieces of about 4×4 mm and culturing themup to day 6. The culture medium was a modified William-E and it has beenchanged at the day 3.

Samples of the sequential extracts as described by Extraction Example 2were air-dried and then redissolved in a quantity of DMSO suitable toobtain a final concentration of 1 and 10 μg/ml. The experimentaltreatments were daily replicated applying 5 ul of extract, solved inpure DMSO, on the surface of the cultured skin samples.

After six days of organ culture, histological section were prepared fromthe skin samples and the quantitative changes of melanin content havebeen investigated following Fontana-Masson staining technique. Themelanin quantification was obtained by image analysis ofmicrophotographs of each histological skin section. Table 10 and FIG. 8summarize the results:

TABLE 10 Melanin content of skin at day 6 of culture - Data from singledonor Melanin content in [%] of the control performance ± standard errorExperimental Melanin Standard N. of ANOVA Ex. Treatment content (%)error Samples Test 0 Control 100 4.8 12 43 dir-Hex 131.1 6.3 12 P < 0.051 μg/ml 44 dir-Hex 147.6 6.3 12 P < 0.01 10 μg/ml 45 seq-EtAc 140.6 8.812 P < 0.05 1 μg/ml 46 seq-EtAc 163.9 15.1 12 P < 0.01 10 μg/ml 47seq-EtOH 129.2 12 12 P < 0.05 1 μg/ml 48 seq-EtOH 149.7 6 12 P < 0.01 10μg/ml 49 seq-Water 153 7.5 12 P < 0.01 1 μg/ml 50 seq-Water 126.3 7.3 12P < 0.05 10 μg/ml

In this experiment all the treatments produced a significant(ANOVA=P<0.05) increase of the skin melanin content. The treatments withdir-Hex 10 μg/ml, seq-EtAc 1-10 μg/ml, seq-EtOH 10 μg/ml and seq-Water 1μg/ml stimulated statistically highly significant responses(ANOVA=P<0.01).

The effects resulted in general in more intense pigmentation than inhair follicles, however, also in this case the more relevant responseshave been recorded by treating the tissue with seq-EtAc (10 μg/ml) andseq-Water (1 μg/ml) extracts.

Effects Examples 51-54: Modulation of Skin Pigmentation by Dir-EtOH

The same protocol described for previous examples 43-50 has been adoptedin the following experiment planned to investigate the activity on skinpigmentation related to dir-EtOH extract according to Extraction Example1.

The extract preparation has been performed using two samples of TahitianIsochrysis biomass, obtained from two different cultivations of the samealgal strain. In order to compare the performance of the two extracts,they have been labelled as dir-EtOH1 and dir-EtOH2. Table 11 and FIG. 9show the results:

TABLE 11 Melanin content of skin at day 6 of culture - Data from singledonor Melanin content in [%] of the control performance ± standard errorExperimental Melanin Standard N. of ANOVA Ex. Treatment content (%)error Samples Test 0 Control 100 10.8 8 51 dir-EtOH1 70.9 9.2 6 1 μg/ml52 dir-EtOH1 67.1 9.1 8 P < 0.05 10 μg/ml 53 dir-EtOH2 57.8 4.4 8 P <0.01 1 μg/ml 54 dir-EtOH2 83.8 12.1 8 10 μg/ml

The results show that both dir-EtOH extracts inhibited the pigmentation,in contrast to the stimulation detected using the extracts obtained withdifferent extractants. Both samples of Tahitian Isochrysis showed thesame effects despite that they originated from different cultures. Allresponses were inhibiting and dir-EtOH1 at 10 μg/ml produced a responsesignificant on statistical basis (P<0.05) while dir-EtOH2 gave a verysignificant response at 1 μg/ml (P<0.01).

This means that different actives are present in Isochrysis sp.,preferably Tahitian Isochrysis, biomass and they can be separated bychoosing the appropriate solvent.

Effects Examples 55-62: Modulation of Skin Pigmentation by Dir-EtOH and“Sequential Extracts”

The same protocol described for previous Effects Examples 51-54 wasadopted in an experiment designed to compare the activity of dir-EtOHwith some “sequential extracts” according to Extraction Example 2. Theaim of the experiment also was to confirm presence of different activesin Tahitian Isochrysis biomass and the possibility to separate them byusing different solvents.

The extracts included in the experiment and obtained results are showedin Table 12 and FIG. 10:

TABLE 12 Melanin content of skin at day 6 of culture - Data from singledonor Melanin content in [%] of the control performance ± standard errorExperimental Melanin Standard N. of ANOVA Ex. Treatment content (%)error Samples Test 0 Control 100 22.3 8 55 dir-EtOH 90.3 11.5 8 1 μg/ml56 dir-EtOH 90.7 10 8 10 μg/ml 57 seq-EtAc 92.7 11.2 8 1 μg/ml 58seq-EtAc 164.9 22.1 8 P < 0.01 10 μg/ml 59 seq-EtOH 131.5 10.5 8 1 μg/ml60 seq-EtOH 157.3 21.6 8 P < 0.05 10 μg/ml 61 seq-Water 138.8 20.5 8 1μg/ml 62 seq-Water 111.2 21 8 10 μg/ml

The results are consistent with the expectations: dir-EtOH tended toinhibit pigmentation, while seq-EtOH produced a significant stimulation(P<0.05). Seq-EtAc (P<0.01) and seq-Water again stimulated the melaninsynthesis as expected (see Effects Examples 45-50).

Effects Examples 63-65: Modulation of Hair Growth by Sequential Extracts

A “sequential extract” was prepared as described in extraction example2. Two extracts were prepared (dir-MeOH followed by seq-Hex/EtAc) and athree dose treatment was tested for the seq-Hex/EtAc extract. Theexperiment has been repeated four times using hair follicles taken from4 different donors and the data have been pooled to represent theaverage response of the four donors. Table 13 summarizes the results ofthe seq-Hex/EtAc extract:

TABLE 13 Growth of hair follicles at the day 9 of culture - Data pooledfrom four replicates Elongation in [%] of the control performance ±standard error Experimental Growth Standard N. of Hair ANOVA Ex.Treatment (%) Error follicles Test 0 Control 100 2.6 62 63 Seq-Hex/EtAc104.4 2.6 43 n.s. 0.1 μg/ml 64 Seq-Hex/EtAc 112.0 3.0 44 P < 0.01 1μg/ml 65 Seq-Hex/EtAc 101.9 3.4 41 n.s. 10 μ/ml

The results attest that the addition of the seq-Hex/EtAc extract leadsto a significant increase in growth of the hair follicles, in comparisonto the untreated group. The most significant response has been obtainedby treating at the dosage of 1 μg/ml which results highly significantalso on a statistical basis (P<0.01).

The seq-Hex/EtAc dry extract only contained trace amounts of PUFAs (<0.2wt. %).

Effects Examples 66-67: Modulation of Skin Pigmentation by “SequentialExtracts”

The same protocol described for previous Effects Examples 51-54 wasadopted in an experiment designed to determine the activity of a“sequential extract” according to Extraction Example 2. Two extractswere prepared (dir-MeOH followed by seq-Hex/Et/Ac) and a three dosetreatment of the seq-Hex/EtAc extract was tested.

The results are shown in Table 14.

TABLE 14 Melanin Content of skin at the day 6 of culture -Data fromsingle donor Melanin content in [%] of the control performance ±standard error Experimental Melanin Standard N. of Ex. Treatment Content(%) Error Samples 0 Control 100 13.7 8 66 Seq-Hex/EtAc 131.3 13.0 8 1μg/ml 67 Seq-Hex/EtAc 110.0 11.4 8 10 μg/ml

In this experiment both treatments produced an increase or melanincontent. The treatment with 1 μg/ml led to an increase of about 30% inmelanin content.

Product Examples 1-11: Skin Care

In table 1 means

-   -   1=Skin tanning “water-in-oil” emulsion    -   2=Skin tanning “oil-in-water” emulsion with UVA/B broadband        protection    -   3=Skin tanning and hair growth inhibiting “oil-in-water” cream    -   4=Hair growth inhibiting aerosol foam with UVB/UVA protection    -   5=Skin lightening and hair growth inhibiting cream O/W    -   6=Skin tanning moisturizing balm    -   7=Skin tanning body spray O/W    -   8=Skin lightening and hair growth inhibiting gel    -   9=Skin tanning and hair growth inhibiting soaking liquid for        wipes    -   10=Hair growth inhibiting and hair lightening antiperspirant        pump spray    -   11=Skin lightening non-aerosol foam

TABLE 1 RAW MATERIAL NAME (SUPPLIER) INCI 1 2 3 4 5 6 7 8 9 10 11 PPMTahitian Isochrysis 10 25 Isochrysis dry Extract extract TahitianIsochryis Maltodextrin. 50 500 100 extract (dry Isochrysis extractcontent 1 Extract wt %) Isochrysis extract Glycerin, Water 500 100 150(dry extract (Aqua), content 0.2 wt %) Pentylene Glycol, IsochrysisExtract WEIGHT % Tahitian 1,2- 10 200 300 Isochrysis Pentylenegly-extract (dry Col, Isochrysis extract content 1 Extract Wt %) Abil 100 ®Dimethicone 2.0 0.5 1.0 (Goldschmidt) Alugel 34 TH Aluminium 1.0(Baerlocher) Stearate alpha-Bisabolol Bisabolol 0.1 0.1 0.2 0.1 0.1 0.10.1 (Symrise) Aristofiex AVC Ammonium 0.8 (Clariant) Acryloyldimethyl-taurate/VP Copolymer Beta-Arbutin Arbutin 0.2 Butylene Glycol ButyleneGlycol 10.0 Caffeine Caffeine 0.2 0.3 Carbopol Carbomer 0.5 Ultrez-10(Noveon) Carbopol Carbomer 0.1 0.1 2050 ® (B.F. Goodrich) Cetiol OEDicaprylyl Ether 3.0 (Cognis) Dracorin GOC Glyceryl Oleate 2.0 (Symrise)Citrate, Caprylic/Capric Triglyceride Drago-Oat- Water (Aqua), 1.0Active Butylene Glycol, Avena Sativa (Oat) Kernel Extract Dragoxat 89Ethylhexyl 5.0 3.0 2.0 (Symrise) Isononanoate Dragoxat EH Ethylhexyl 3.03.0 (Symrise) Ethylhexanoate Edeta BD ® Dinatrium-EDTA 0.1 0.1 0.1 0.10.1 (BASF) Emulgade PL Cetearyl 0.5 Glucoside, Cetearyl AlcoholEmulsiphos Cetylphosphate, 2.0 1.5 (Symrise) Hydrogenated Palmglycerides Ethanol (96 %) Alcohol Denat. 2.0 60.0 Extrapone AloeGlyerin, Water 3.0 0.5 Vera (Symrise) (Aqua), Aloe Barbadensis LeafExtract Extrapone Glycerin, Water 0.5 Chamomile (Aqua), (Symrise)Chamomilla Recutica (Matricaria), Flower, Extract Extrapone Glycerin,Water 0.2 Green Tea (Aqua), Cemallia (Symrise) Sinensis Leaf ExtractExtrapone Glycerin, Water 0.3 Rosemary (Aqua), (Symrise) RosmarinusOfficinalis (Rosemary) Leaf Extract Extrapone Propylene Glycol, 1.0Witch Hazel Hamamelis (Symrise) Virginiana (Witch Hazel) Water, Water(Aqua), Hamamelis Virginiana (Witch Hazel) Extract Fragrance Parfum 0.40.5 0.3 0.4 0.3 0.3 0.2 0.2 0.5 1.0 0.2 (Fragrance) Glycerin 99%Glycerin 2.0 4.0 2.0 3.0 1.5 5.0 4.0 Hostacerin Polyglcery1-2- 3.0DGMS ® Stearate (Clariant) Hydrolite-5 1,2- 3.5 5.0 5.0 1.0 5.0 5.0(Symrise) Pentyleneglycol Hydroviton 24 Water (Aqua), 1.0 (Symrise)Pentylene Gylcol, Glycerin, Sodium Lactate, Lactic Acid, Serine, Urea,Sorbitol, Sodium Chloride, Allantoin Isodragol Triisononanoin 2.0(Symrise) Isopropylmyristat Isopropyl Myristate 4.0 (Symrise) Karion FSorbitol 2.0 (Merck) Keltrol T ® Xanthan Gum 0.2 0.1 0.3 (Calgon) Kojicacid Kojic acid 0.2 Lanette E ® Natriumcetearyl- 0.7 (Cognis) sulfatatLanette O ® Cetearyl Alcohol 3.0 (Cognis) Lanette 16 ® Cetyl alcohol 2.00.5 1.0 (Cognis) Lara Care A- Galactoarabinan 0.2 200 (Rahn) Locron LAluminium 16.0 (Cognis) Chlorohydrate Magnesium Magnesium Sulfate 0.7Sulfat Hepathydrat (Merck) Mineral Oil Paraffinum 4.0 Liquidum NaOH 10%Sodium hydroxide 0.2 2.9 0.4 1.0 0.6 aq. solution Neo Heliopan ®Dinatrium- 6.7 AP Phenyldibenzimida- (Symrise), 15 zoltetrasulfonate %as sodium salt Neo Heliopan ® Ethylhexylmethoxy- 6.0 2.0 AV (Symrise)cinnamate Neo Heliopan ® Benzophenone-3 0.25 BB (Symrise) Neo Heliopan ®Butyl 0.6 1.5 1.5 357 (Symrise) Methoxydibenzoyl- methane Neo Heliopan ®Isoamyl-p- 6.0 E 1000 methoxycinnamate (Symrise) Neo Heliopan ®Homosalate 9.5 HMS (Symrise) Neo Heliopan ® Phenylbenzimidazol 6.7 13.3Hydro sulfonic acid (15% aq. solution neutralized with NaOH) (Symrise)Neo Heliopan ® 4- 4.0 3.0 MBC Methylbenzyliden- (Symrise) campherNeo-PCL Trideceth-9, PEG-5 1.0 2.0 Water Soluble Ethylhexanoate, N(Symrise) Water (Aqua) Neutral oil Caprylic/Capric 5.0 0.25 2.0 6.0 4.0(Symrise) Triglyceride PCL liquid 100 Cetearyl 12.0 5.0 3.0 3.0 7.0(Symrise) Ethylhexanoate PCL solid Stearyl 2.0 1.5 (Symrise) Heptanoate,Stearyl Caprylate Pemulen TR-2 Acrylates/C10-30 0.2 0.2 (Noveon) AlkylAcrylate Crosspolymer Phenoxyethanol Phenoxyethanol 0.7 0.7 0.7(Symrise) Prisorine Isostearic acid 0.5 3505 ® (UniQema) 1,2- PropyleneGlycol 5.0 5.0 3.0 Propylenglycol Sepigel 305 Polyacrylamide, 1.0 C13-14Isoparaffin, Laureth-7 SF1214 ® Cyclopentasiloxane, 1.0 (Bayer)Dimethicone Solubilizer PEG-40 1.0 1.75 3.0 (Symrise) HydrogenatedCastor Oil, Trideceth-9, Propylene Glycol, Water (Aqua) Sun Flower OilHelianthus Annus 5.0 (H. Erhard (Sunflower) Seed Wagner) Oil Sweet AlmonPrunus Dulcis 5.0 Oil (H. Erhard Wagner) SymDeo MPP Dimethyl Phenyl 2-0.5 (Symrise) Butanol Symdiol 68 1,2-Hexanediol, 0.5 0.5 (Symrise)Caprylalcohol SymGlucan Water, Glycerin, 0.3 (Symrise) Beta-GlucanSymWhite 377 Phenylethyl 0.2 (Symrise) Resorcinol Tegosoft TN ® C12-C152.0 2.0 (Goldschmidt) Alkylbenzoate Texapon N 70 Sodium Laureth 0.1 0.50.5 (Cognis) Sulfate Vitamin E Tocopheryl Acetate 3.0 0.5 0.5 0.5Acetate (DSM Nutritional Products) Vitamin A Retinyl Palmitate 0.2Palmitate in oil (1 Mio le/g) (DSM Nutritional Products) Water, Aqua(Water) ad ad ad ad ad ad ad ad ad ad ad demineralized 100 100 100 100100 100 100 100 100 100 100

Product Examples 12-20: Hair Care

In table 2 means

12=Hair growth stimulating tonic13=2 in 1 After sun shampoo with hair tanning properties14=Hair tanning conditioner with UVB/UVA protection15=Liquid hair leave-on, pump-foam with hair growth stimulatingproperties16=Hair growth stimulating styling gel17=Hair growth inhibiting and hair lightening setting foam18=Mascara with hair growth stimulating and hair tanning properties19=Hair growth stimulating anti-dandruff shampoo20=Hair growth inhibiting leave-on hair conditioner21=Hair tanning and hair growth stimulating shampoo Table 2:

RAW MATERIAL NAME (SUPPLIER) INCI 12 13 14 15 16 17 18 19 20 21 PPMTahitian Isochrysis 25.0 10.0 Isochrysis dry Extract extract Isochryisextract Maltodextrin, 20 100 (dry extract Isochrysis content 0.5 wt %)Extract Isochrysis extract Propylene 25 500 150 (dry extract Glycol,Water content 2 wt %) (Aqua), PEG-40 Hydrogenated Castor Oil,Trideceth-9, Isochrysis Extract Tahitian Isochryis 1,2- 100 200 500extract (dry Pentylenegly- extract content 1 Col, Water wt %) (Aqua),Isochrysis Extract WEIGHT % Abil B 9950 Dimethicone 0.2 (Evonic PropylPg- Goldschmidt) Betaine Abil-Quat 3272 Quaternium-80 0.5 (EvonicGoldschmidt) Actipone Water 0.75 AlphaPulp (Aqua), Butylene (Symrise)Glycol, Malic Acid, Actinidia Chinensis (Kiwi) Fruit Juice, CitrusAurantium Dulcis (Orange) Juice, Citrus Paradisi (Grapefruit) Juice,Pyrus Malus (Apple) Juice, Trideceth- 9, Prunus Amygdalus Dulcis (SweetAlmond) Seed Extract Aloe Vera Gel Water (Aqua), 0.5 0.5 Concentrate10/1 Aloe (Symrise) Barbadensis Leaf Juice -(-Alpha-)- Bisabolol 0.1Bisabolol, natural (Symrise) Aminexil Diaminopyrimidine 0.3 Oxide Antil141 Liquid Propylene 1.0 (Evonic Glycol, PEG-55 Goldschmidt) PropyleneGlycol Distearate Antil 171 (Evonic PEG-18 Glyceryl 2.0 Goldschmidt)Oleate/Cocoate Caffeine Caffeine 0.5 0.1 0.3 Carbopol Ultrez- Carbomer0.7 110 (Noveon) Celquat L-200 Polyquaternium- 1.0 (National Starch 4 &Chemical) CeramideBio N-(1- 0.2 Hexadecanoyl)- 4-hydroxy-L- prolin-(1-hexadecyl-ester Citric Acid 10% Citric Acid 1.3 1.3 q.s. solution Colour(Symrise) Colour 0.2 Crinipan AD Climbazole 0.5 (Symrise) Crotein Q(Croda) Hydroxypropyl 1.0 Trimonium Hydrolyzed Collagen Dehyquart A CACetrimonium 0.2 1.0 4.0 4.0 (Cognis) Chloride Dehyquart SP Quaternium-520.5 (Cognis) Dehyton K Cocamidopropyl 8.0 0.5 2.0 (Cognis) BetaineDermosaccharides Water (Aqua), 1.0 GY (Impag) Glycerin, GlycogenD-Panthenol 75L Panthenol 0.5 1.0 0.5 (DSM Nutritional) Dow Corning 245Cyclopentasiloxane 5.0 Fluid Dow Corning Cyclopentasiloxane, 1.0 5225CPEG/PPG- Formulation Aid 18/18 Dimethicone Dracorin GMS Glyceryl 2.0(Symrise) Stearate Dragocide Liquid Phenoxyethanol, 0.8 0.5 0.5 0.7 0.80.5 (Symrise) Methyl-, Ethyl-, Butyl-, Propyl-, IsobutylparabenDragocolor Blue Basic Blue 99 0.02 (Symrise) Dragocolor Basic Brown 170.1 Brown (Symrise) Dragocolor Basic Brown 16 0.1 Mahagony (Symrise)Dragoderm Glycerin, 1.0 2.0 (Symrise) Triticum Vulgare (Wheat) Gluten,Water (Aqua) Edeta BD (BASF) Disodium EDTA 0.05 Emulgin B2 Ceteareth-200.7 (Cognis) Ethanol 96% Ethanol 48.0 3.0 5.0 13.0 Euperlan PK 771Glycol 3.0 (Cognis) Distearate, Sodium Laureth Sulfate, Cocamide MEA,Laureth-10 Euperlan PK 900 PEG-3 Distearate 2.0 BENZ-W (Cognis) EuperlanPK 4000 Glycol Distearate, 2.5 (Cognis) Laureth-4, Cocoamidopro-pylBetaine Ewacera 12 (H, Bees Wax 10.0 Erhard Wagner) Ewacera 34 (H.Carnauba Wax 4.0 Erhard Wagner) Extrapone Water (Aqua), 0.5 CamomilePropylene Glycol, (Symrise) Butylene Glycol, Chamomilla Recutita(Mathcaria) Flower Extract, Glucose, Bisabolol Extrapone Green Glycerin,Water 0.3 Tea GW (Symrise) (Aqua), Camellia Sinensis Leaf ExtractExtrapone Hop GWT Glycerin, Water 0.4 (Aqua), Humulus Lupulus (Hops)Cone Extract, Glucose Extrapone Propylene Glycol, 0.4 1.0 LemongrassWater (Aqua), (Symrise) PEG-40 Hydrogenated Castor Oil, Trideceth-9,Cymbopogon Citratus Leaf Oil, Lactic Acid Extrapone Rosemary Glycerin,Water 0.3 GW (Symrise) (Aqua), Rosmarinus officinalis (Rosemary) LeafExtract Fragrance (Symrise) Fragrance 0.5 0.3 0.4 0.2 0.4 0.5 0.5 0.30.4 Frescolat ML Menthyl Lactate 0.5 0.8 0.5 (Symrise) Glycerin, 99.5%Glycerin 10.0 Hydrolite-5 Pentylene Glycol 0.5 (Symrise) Keltrol T(Calgon) Xanthan Gum 0.15 Lanette 18 (Cognis) Stearyl Alcohol 2.0Lanette O (Cognis) Cetearyl Alcohol 2.5 3.5 Luviskol K 30 (BASF) PVP 2.0Luviskol K 30 PVP/ 3.0 4.0 Powder Polyvinylpyrrolidone (BASF) LuviskolVA 64 PVP/VA Copolymer 4.0 Powder (BASF) MBD 210 20% Carbon Black, 5.0Dispersion Water (Aqua) Minoxidil Minoxidil 0.5 Merquat 550Polyquaterinium-7 1.0 (Ondeo) Mulsifan RT 203/80 C12-15 Pareth-12 4.0(Z&S) Neo Heliopan Butyl 0.5 357 (Symrise) Methoxydibenzo- ylmethane NeoHeliopan BB Benzophenone-3 0.1 0.2 0.3 (Symrise) Neo Heliopan EIsoamyl-p- 2.0 1000 (Symrise) methoxy- cinnamate Neo-PCL WaterTrideceth-9, 1.0 soluble N PEG-5 (Symrise) Ethylhexanoate, Water (Aqua)Neutrol TE Tetrahydroxypropyl 1.4 (BASF) Ethylendiamine NiacinamideNiacinamide 0.2 Permethyl 104A Polyisobutene 1.0 (Cesham) C68 Plantacare1200 Lauryl glucoside 10.0 UP (Cognis) Polymer JR 400 Polyquaternium-0.2 (Nordmann, 10 Rassmann) Polyquart H 81 PEG-15 Coco 3.0 (Cognis)Polyamine Potassium Potassium 0.2 Sorbate Sorbate Rose CL forte Water(Aqua), 0.5 (Symrise) Glycerin, PEG-40 Hydrogenated Castor Oil, RosaDamascena Flower Oil Seanamin FP LS Hydrolyzed Actin, 1.0 5988 (Impag)Water (Aqua), Glycerin, Fucus Vesiculosus Extract Sodium Benzoate SodiumBenzoate 0.5 Sodium Chloride Sodium Chloride 0.5 2.0 2.0 SodiumHydroxide, Sodium Hydroxide 0.1 10% sol. Solubilizer PEG-40 1.0(Symrise) Hydrogenated Castor Oil, Trideceth-9, Water (Aqua) StearicAcid Stearic Acid 2.0 (Cognis) Tego Betain 810 Capryl/Capramido 0.5(Evonic propyl Betaine Goldschmidt) Texapon K 14 S Sodium Myreth 12.0Special (Cognis) Sulfate Texapon N 70 Sodium Laureth 10.0 12.0 (Cognis)Sulfate Triethanolamine Triethanolamine 0.5 Veegum HV Magnesium 0.55Aluminium Silicate Water, Wasser (Aqua) Ad Ad Ad Ad Ad Ad Ad Ad Ad ADdemineralized 100 100 100 100 100 100 100 100 100 100

Product Examples 22-34: Beauty from Inside

Gelatine capsule for direct consumption

RAW MATERIAL WEIGHT % NAME 22 23 24 Gelatine shell: Glycerin 2.014 2.0142.014 Gelatine 240 Bloom 7.91 7.91 7.91 Sucralose 0.065 0.065 0.065Allura Red 0.006 0.006 0.006 Brillant Blue 0.005 0.005 0.005 Corecomposition: Plant oil triglyceride ad 100 ad 100 ad 100 (coconut oilfraktion) Aroma B 9.95 12.0 12.0 Tahitian Isochrysis dry 0.005 0.01extract Isochrysis extract (plant 0.02 oil triglyceride:dry extract 95:5(w/w)

Aroma B had the following composition (figures in wt. %): 0.1% neotamepowder, 0.05% aspartame, 29.3% peppermint oil arvensis, 29.3% peppermintpiperita oil Willamette, 2.97% sucralose, 2.28% triacetin, 5.4% diethyltartrate, 12.1% peppermint oil yakima, 0.7% ethanol, 3.36%2-hydroxyethyl menthyl carbonate, 3.0% 2-hydroxypropyl menthylcarbonate, 0.27% vanillin, 5.5% D-limonene, 5.67% L-menthyl acetate.

The gelatine capsule suitable for direct consumption (produced in ananalogous way to WO 2004/050069) had a diameter of 5 mm and the weightratio of core material to shell material was 90:10. The capsule openedin the mouth in less than 10 seconds and dissolved completely in lessthan 50 seconds.

Compressed tablets

RAW MATERIAL WEIGHT % NAME 25 26 27 Magnesium stearate 0.9 0.9 0.9 (aslubricant) Citric acid 0.2 0.2 0.2 Isochrysis dry extract 0.01 0.0050.001 Dextrose Ad 100 Ad 100 Ad 100

Preparation instructions: Mix all the constituents and press to acompressed product in a suitable machine.

Chewing gums

RAW MATERIAL WEIGHT % NAME 28 29 Chewing gum base 21.0 30.0 Glycerin 0.51.0 Menthol spearmint 1.0 0.7 aroma Glucose syrup 16.5 Icing sugar Ad100 Tahitian Isochrysis dry 0.01 extract Isochryis extract 0.5(maltodextrimdry extract 99:1 (w/w)) Sorbitol, powdered Ad 100Palatinite 9.5 Xylitol 2.0 Mannitol 3.0 Aspartame 0.1 Acesulfame K 0.1Emulgum/emulsifier 0.3 Sorbitol 70%, in water 14.0

In table 3 means

30=Instant beverage mix31=Sugar-free instant beverage mix32=Carbonated soft drink33=Soja-fruit drink34=Low-fat yoghurt

TABLE 3 RAW MATERIAL WEIGHT % NAME 30 31 32* 33 34 Tahitian Isochrysis0.005 0.007 0.01 dry extract Isochryis extract 0.2 0.02(maltodextrin:dry extract 95:5 (w/w)) Sugar (sucrose) ad 100 Citric acid4.00 33.33 0.2 Trisodium citrate 0.26 Tricalcium phosphate 0.22 Ascorbicacid 0.24 0.44 (vitamin C) Clouding agent and 0.20 titanium dioxide (E171) Xanthan gum (E 415) 0.072 Sodium carboxy 0.064 methyl cellulose (E467) Pectin (E 440) 0.04 Spray-dried pineapple 0.40 flavor, includingyellow colorant tartrazine Spray-dried raspberry 11.50 flavor, includingred colorant Lemon and lime 0.01 flavor D-Limonene 0.005 Maltodextrin(powder) ad 100 Aspartame 3.30 Saccharose 8.0 6.0 5.0 Hesperetin (1 wt.% in 0.05 1,2-propylene glycol) Phloretin (1 wt. % in 0.05 1,2-propyleneglycol) Ethylhydroxymethyl- 0.01 ppb  furanone Vanilla flavor 0.10 0.125Vanillin  15 ppb Maltol 350 ppb 2,5-Dimethyl-4-  3 ppbhydroxy-2H-furan-3- one 1,2-Propylene glycol 0.1 Mixture of fruit juice45.0 concentrates Soja powder 5.0 Yoghurt (1.5 wt. % ad 100 fat) Waterad 100 ad 100 *Carbonated after filling into bottles.

It is claimed:
 1. A method of obtaining a composition for influencing ormodifying a) growth of human hair and/or b) pigmentation of human skinand/or hair, comprising the step of extracting cell material ofIsochrysis sp. with a liquid extractant selected from the groupconsisting of hexane, ethyl acetate, ethanol, water, methanol,isopropanol and mixtures of two or more of these extractants, whereinthe extraction comprises a) exposition of the cell material to theextractant for up to 24 h at a temperature of not more than 50° C., andb) removal of the cell material to obtain an extract, the extract beingthe composition or being further processed into the composition, andwherein the cell material of Isochrysis sp. is obtainable or obtained bya method consisting of the steps:
 1. Cultivating Isochrysis sp. cells,2. Harvesting the cells to obtain completely or substantially intactcell material,
 3. Optionally washing the cell material of step 2 once ormultiple times, to obtain washed, substantially or completely intactcell material,
 4. Optionally freeze-drying the cell material of step 2and/or step
 3. 2. The method according to claim 1, wherein theextraction further comprises repeating steps a) and b) once, twice,three or four times, and wherein in each step a) the cell materialremoved in the respective prior step b) is used, and the extracts ofsteps b) are combined.
 3. The method according to claim 1, furthercomprising the step of adjusting the phototoxicity of the composition toa phototoxicity index of less than
 5. 4. The method according to claim3, wherein for adjusting the phototoxicity the extract is treated withactivated carbon in a ratio dry extract:activated carbon of 3:1 to 1:20,all weights given as dry weights.
 5. The method according to claim 1,wherein for extraction cell material is obtained in step b) of a priorextraction with a different extractant.
 6. The method according to claim1, wherein the composition comprises 0.001 to 20 wt. % dry extract ofthe total composition.
 7. Method according to claim 1, wherein the cellmaterial used in the first step a) is freeze-dried Isochrysis sp.
 8. Acomposition for influencing or modifying a) growth of human hair and/orb) pigmentation of human hair and/or c) pigmentation of human skin, saidcomposition being obtainable or obtained by a method according toclaim
 1. 9. A composition according to claim 8 for stimulating growth ofhuman hair without increasing pigmentation of human skin and/or hair,characterized in that the composition is or comprises an extractobtained by using methanol as extractant, or for stimulating growth ofhuman hair and increasing pigmentation of human skin and/or hair,characterized in that the composition is or comprises an extractobtained by using ethyl acetate as extractant or by using a mixture ofhexane/ethyl acetate as extractant on cell material obtained afterextraction with methanol, or for increasing pigmentation of human skinand/or hair without stimulating growth of human hair, characterized inthat the composition is or comprises an extract obtained by using wateras extractant, or for inhibiting growth of human hair while increasingpigmentation of human skin and/or hair, characterized in that thecomposition is or comprises an extract obtained by using ethanol asextractant on cell material obtained after extraction with ethylacetateor with hexane followed by extraction with ethylacetate, or forinhibiting growth of human hair and decreasing pigmentation of humanskin and/or hair, characterized in that the composition is or comprisesan extract obtained by using ethanol as extractant.
 10. A cosmetic,dermatological or therapeutic product comprising a composition accordingto claim 8, wherein the product is a skin care composition,hair-removing composition, hair care composition, decorative cosmetic,or composition for oral application.
 11. A method for influencing ormodifying a) growth of human hair and/or b) pigmentation of human hairand/or c) pigmentation of human skin, the method comprising applying acomposition being or comprising an extract of Tahitian Isochrysis,wherein the extract is obtainable or obtained by a method according toclaim
 1. 12. The method according to claim 11, wherein the method is forstimulating growth of human hair without increasing pigmentation ofhuman skin and/or hair, for stimulating growth of human hair andincreasing pigmentation of human skin and/or hair, for stimulatinggrowth of human hair and decreasing pigmentation of human skin and/orhair, for increasing pigmentation of human skin and/or hair withoutstimulating growth of human hair, for inhibiting growth of human hairand increasing pigmentation of human skin and/or hair, for inhibitinggrowth of human hair without increasing pigmentation of human skinand/or hair, or for inhibiting growth of human hair and decreasingpigmentation of human skin and/or hair.
 13. The method of claim 1,wherein the Isochrysis sp. is Tahitian Isochrysis.
 14. The methodaccording to claim 3, wherein for adjusting the phototoxicity theextract is treated with activated carbon in a ratio dryextract:activated carbon of 1:1 to 1:12, all weights given as dryweights.
 15. The method according to claim 1, wherein the compositioncomprises 0.01 to 10 wt % dry extract of the total composition.
 16. Themethod according to claim 1, wherein the composition comprises 0.1-5 wt% dry extract of the total composition.
 17. The cosmetic, dermatologicalor therapeutic product of claim 10, wherein the skin care composition isan emulsion, ointment, paste, gel, alcoholic or aqueous/alcoholicsolution, oil, toner, balsam, serum, powder, or soaking liquid forwipes.
 18. The cosmetic, dermatological or therapeutic product of claim10, wherein the skin care composition has a presentation form as a mask,mousse, stick, pencil, roll-on, spray, or aerosol.
 19. The cosmetic,dermatological or therapeutic product of claim 10, wherein the skin carecomposition is a sunscreen composition, self-tanning composition and/oraftersun preparation, shaving composition or after-shave, or deodorantand/or antiperspirant.
 20. The cosmetic, dermatological or therapeuticproduct of claim 10, wherein the hair care composition is a shampoo,conditioner, hair treatment cure, hair tonic, hair lotion, hair rinse,styling cream, hair setting composition, styling aid, blondingcomposition, hair colouring composition, or decorative cosmetic.
 21. Acosmetic, dermatological or therapeutic product comprising a compositionaccording to claim 10, wherein the product is a composition for oralapplication.
 22. The cosmetic, dermatological or therapeutic productaccording to claim 21, wherein the composition for oral application isin the form of tablets, dragees, capsules, juices, solutions, granulesand foodstuff.